Cosmetic Composition and Preparation Method and Use Thereof

ABSTRACT

A skin-whitening, freckle-removing cosmetic composition, comprising an effective amount of extract from  Cistanche deserticola  and other useful adjuvants.

FIELD OF THE INVENTION

The present invention relates generally to a cosmetic compositioncomprising an extract that is derived from a plant, a process forpreparing same and a use thereof. More particularly, the inventionrelates to a dermatologic and cosmetic preparation comprising an extractderived from a plant, which possesses skin-whitening andfreckle-removing actions and makes the skin more youthful looking. Thepresent invention also relates to a process for preparing saidpreparation.

BACKGROUND OF THE INVENTION

With the elevation of living standard, women of all ages pay more andmore attention to the skin health, and maintaining a youthful lookingskin becomes an on-going need for women. To improve the youthfulappearance of skin generally includes the improvement of overall skintone, reduction in pigmentation and spots, and increases in supplenessand softness. Genus cistanche is a traditional Chinese medicine ofkidney-invigoration, having pneuma-tonifing and antidotal actions. Itshistory as both a drug and a food for the double purpose is over twothousand years.

Japan scholars have recently found that phenylethanoid glycosides, asactive components obtained from plants, are of more potenttyrosinase-inhibiting activity and chromatogenesis-inhibiting activity.It has been shown, in vitro, that phenylethanoid glycosides possess anactive promotion on cytolergy of M cells to form a collagen fibernetwork (Tanimoto S, et al., Yakugaku Zasshi. 2006 March; 126(3):173-7.Sudo H et all. Chem Pharm Bull (Tokyo). 1999 September; 47(9):1341-3.Related).

It has been found that the extract from Genus cistanche containsphenylethanoid glycosides. Among all species of the genus cistanche,Cistanche tubulosa (Schenk.) Wight produced in southern Xinjiang, China,contains the greatest amount of phenylethanoid glycosides, thus isexpectable to be used in skin care preparations to improve skin healthand produce agerasia and facial actions.

SUMMARY OF THE INVENTION

The primary object of the present invention is to provide a cosmeticcomposition comprising an extract obtained from Genus cistanche. It isanother object of the present invention to provide a process forpreparing an extract which is rich in phenylethanoid glycosides fromCistanche tubulosa (Schenk.) wight. It is still another object of thepresent invention to provide the application of the cosmetic compositioncontaining such extract of Cistanche tubulosa (Schenk.) Wight.

For the above objects, the solutions of the present invention are asfollows: A cosmetic composition comprising an effective amount ofextract of Cistanche tubulosa (Schenk.) Wight and carrier(s) acceptablein cosmetic.

The extract of Cistanche tubulosa, especially, the extract fromCistanche tubulosa (Schenk.) Wight, is rich in phenylethanoidglycosides. Preferably, the extract comprises at least 2% of acteosideby weight, and at least 4% of echinaoside by weight, based on the totalweight of the extract. As main active ingredients, the higher theamounts of acteoside and echinaoside are, the better the effect is.However, in view of various factors such as the commercial cost, it isimpossible to make unlimited refines to raise the contents of these twoactive ingredients. Therefore, once the content of acteoside reaches2-80% by weight and the content of echinaoside reaches 4-80% by weight,the effect is desirable. Most preferably, the weight radio of acteosideto echinaoside is over 2:1.

Besides that the extract from Cistanche tubulosa (Schenk.) wight is richin phenylethanoid glycosides, the extract also comprises acteosideisomer, 2′acetylacteoside and tubuloside A, B, C, each being in anamount less than 4% by weight based on the total weight of the extract.The contents of acteoside and echinaoside are measured by HPLC, a methodof content determination recorded in page 90 of China Pharmacopeia I(Ed. 2005).

Phenylethanoid glycosides have general formula of:

Ingredient R1 R2 R3 R4 R5 R6 R7 Acteoside H RHa Cf H H OH OHEchinacoside H RHa Cf Glc H OH OH 2′acetylacteoside Ac RHa Cf H H OH OHacteoside isomer H RHa H Cf H OH OH tubuloside A H RHa Cf Glc H H OHtubuloside B H RHa Cm Glc H OH OH tubuloside C H RHa Cm Glc H OH OH

The carrier(s) used in the cosmetic compositions of the presentinvention is/are dermatologically acceptable carrier(s) that is/aresuitable for topical application to the keratinous tissue, andcompatible with the active ingredients of the present invention and anyother additives, without any concern of untoward safety or toxicity tothe skin. A safe and effective amount of carrier(s) is from about 10% toabout 99.99%, preferably from about 20% to about 99.9%, more preferablyfrom about 40% to about 98%, based on the total weight of thecomposition.

The carrier can be in a wide variety of forms. For example, emulsioncarriers, including, but not limited to, oil-in-water, water-in-oil,water-in-oil-in-water (W/O/W), and oil-in-water-in-silicone (W/O/S)emulsions, are useful herein.

Preferred carriers contain an emulsion such as oil-in-water emulsions,water-in-oil emulsions, and water-in-silicone emulsions. As will beunderstood by the skilled artisan, a given component will distributeprimarily into either the water or oil/silicone phase, depending on thewater solubility and dispersibility of the component in the composition.

Emulsions according to the present invention generally contain Lipidsand oils which may be derived from animals, plants or petroleum and maybe natural or synthetic. Preferred emulsions also contain a humectant,such as glycerin or propylene glycol or butylenes glycol or sorbitol.Emulsions will preferably further contain from about 0.01% to about 10%,more preferably from about 0.1% to about 5%, of an emulsifier, based onthe weight of the carrier. Emulsifiers may be nonionic, anionic orcationic. Suitable emulsifiers are disclosed in, for example, U.S. Pat.No. 3,755,560, and McCutcheon's Detergents and Emulsifiers, NorthAmerican Edition, pages 317-324 (1986).

The emulsion may also contain an anti-foaming agent to minimize foaming.Anti-foaming agents include high molecular weight silicones and othermaterials well known in the art for such use.

Water-in-silicone emulsions contain a continuous silicone phase and adispersed aqueous phase.

The organopolysiloxane oil for use in the emulsion may be volatile,non-volatile, or a mixture of volatile and non-volatile silicones. Theterm “nonvolatile” as used in this context refers to those siliconesthat are liquid under ambient conditions and have a flash point (underone atmospheric of pressure) of greater than 100 degree Centigrade. Theterm “volatile” as used in this context refers to all other siliconeoils. Suitable organopolysiloxanes can be selected from a wide varietyof silicones spanning a broad range of volatilities and viscosities.Examples of suitable organopolysiloxane oils include polyalkylsiloxanes,cyclic polyalkylsiloxanes and polyalkylarylsiloxanes.

Polyalkylsiloxanes useful in the composition herein includepolyalkylsiloxanes with viscosities of from about 0.5 to about 1,000,000centistokes at 25° C. Such polyalkylsiloxanes can be represented by thegeneral formula R₃ SiO[R₂SiO]_(x) SiR₃ wherein R is an alkyl grouphaving from one to about 30 carbon atoms (preferably R is methyl orethyl, more preferably ethyl), and x is an integer from 0 to about10,000, chosen to achieve the desired molecular weight. Commerciallyavailable polyalkylsiloxanes include the polydimethylsiloxanes, examplesof which include the Vicasil® series sold by General Electric Companyand the Dow Corning® 200 series sold by Dow Corning Corporation.Specific examples of suitable polydimethylsiloxanes include Dow Corning®200 fluid having a viscosity of 0.65 centistokes and a boiling point of100° C., Dow Corning® 225 fluid having a viscosity of 10 centistokes anda boiling point greater than 200° C., and Dow Corning® 200 fluids havingviscosities of 50, 350, and 12,500 centistokes, respectively, andboiling points greater than 200° C. Suitable dimethicones include thoserepresented by the formula (CH₃)₃SiO[(CH₃)₂SiO]_(x)[CH₃RSiO]_(y)Si(CH₃)₃wherein R is a straight or branched chain alkyl having from two to about30 carbon atoms and x and y are each integers of 1 to about 10,000,000selected to achieve the desired molecular weight. Examples of thesealkyl-substituted dimethicones include cetyl dimethicone and lauryldimethicone.

Cyclic polyalkylsiloxanes suitable for use in the composition ascarrier(s) include those represented by the general chemical formula[SiR₂—O]_(n) wherein R is an alkyl group (preferably R is methyl orethyl, more preferably methyl) and n is an integer from about 3 to about8, more preferably n is an integer from about 3 to about 7, and stillmore preferably n is an integer from about 4 to about 6. When R ismethyl, these materials are typically referred to as cyclomethicones.Commercially available cyclomethicones include Dow Corning® 244 fluidhaving a viscosity of 2.5 centistokes, and a boiling point of 172° C.,which primarily contains the cyclomethicone tetramer (i.e. n=4), DowCorning® 344 fluid having a viscosity of 2.5 centistokes and a boilingpoint of 178° C., which primarily contains the cyclomethicone pentamer(i.e. n=5), Dow Corning® 245 fluid having a viscosity of 4.2 centistokesand a boiling point of 205° C., which primarily contains a mixture ofthe cyclomethicone tetramer and pentamer (i.e. n=4 and 5), and DowCorning® 345 fluid having a viscosity of 4.5 centistokes and a boilingpoint of 217° C., which primarily contains a mixture of thecyclomethicone tetramer, pentamer, and hexamer (i.e. n=4, 5, and 6).

Also useful are materials such as trimethylsiloxysilicate, which is apolymeric material corresponding to the general chemical formula [(CH₂)₃SiO_(1/2)]_(x) [SiO₂]_(y), wherein x is an integer from about 1 to about500 and y is an integer from about 1 to about 500. A commerciallyavailable trimethylsiloxysilicate is sold as a mixture with dimethiconeand Dow Corning® 593 fluid.

Dimethiconols are also suitable for use in the cosmetic composition.These compounds can be represented by the general chemical formulas R₃SiO[R₂SiO]_(x)SiR₂OH and HOR₂SiO[R₂SiO]_(x)SiR₂OH wherein R is an alkylgroup (preferably R is methyl or ethyl, more preferably methyl) and x isan integer from 0 to about 500, chosen to achieve the desired molecularweight. Commercially available dimethiconols are typically sold asmixtures with dimethicone and cyclomethicone (e.g. Dow Corning® 1401,1402, and 1403 fluids).

Polyalkylaryl siloxanes are also suitable for use in the cosmeticcomposition. Polymethylphenyl siloxanes having viscosities from about 15to about 65 centistokes at 25° C. are especially useful.

Preferred for use herein are organopolysiloxanes selected from any oneof polyalkylsiloxanes, alkyl substituted dimethicones, cyclomethicones,trimethylsiloxysilicates, dimethiconols, polyalkylaryl siloxanes, andany combination thereof. Preferred among the polyalkylsiloxanes aredimethicones.

The cosmetic composition may additionally contain one or morenon-silicone oils. Suitable non-silicone oils have a melting point ofabout 25° C. or less. Examples of such non-silicone oils are those wellknown in the chemical arts in the form of water-in-oil emulsions, e.g.,mineral oil, vegetable oils, synthetic lipids, and semisynthetic lipids,etc.

Dispersed Aqueous Phase when Water in Silicone Emulsion is Used.

The cosmetic compositions of the present invention contain from about10% to about 90%, more preferably from about 20% to about 85%, and stillmore preferably from about 30% to about 80% of a dispersed aqueousphase. The dispersed phase is known as the internal phase ordiscontinuous phase. The dispersed aqueous phase is a dispersion ofsmall aqueous particles or droplets suspended in and surrounded by thecontinuous silicone phase described hereinbefore.

The aqueous phase can be water, or a combination of water and one ormore water soluble or dispersible ingredients. Nonlimiting examples ofsuch ingredients include thickeners, acids, bases, salts, chelants,gums, water-soluble or dispersible alcohols and polyols, buffers,preservatives, sunscreening agents, colorings, and the like.

The cosmetic compositions of the present invention will typicallycontain from about 25% to about 90%, preferably from about 40% to about80%, more preferably from about 60% to about 80%, water in the dispersedaqueous phase by weight of the composition.

Emulsifier for Dispersing the Aqueous Phase

The water-in-silicone emulsions of the present invention preferablycontain an emulsifier. In a preferred embodiment, the compositioncontains from about 0.1% to about 10% emulsifier, more preferably fromabout 0.5% to about 7.5%, still more preferably from about 1% to about5%, emulsifier by weight of the composition. The emulsifier helpsdisperse and suspend the aqueous phase within the continuous siliconephase.

A wide variety of emulsifying agents can be employed herein to form thewater-in-silicone emulsion. Known or conventional emulsifying agents canbe used in the present invention, provided that the selected emulsifyingagent is chemically and physically compatible with components of thecomposition of the present invention, and provides the desireddispersion characteristics. Suitable emulsifiers include siliconeemulsifiers, non-silicon-containing emulsifiers, and mixtures thereof,known by those skilled in the art for use in topical personal careproducts. Preferably these emulsifiers have an HLB value(hydrophile-lipophile balance value) of or less than about 14, morepreferably from about 2 to about 14, and still more preferably fromabout 4 to about 14. Emulsifiers having an HLB value outside of theseranges can be used in combination with other emulsifiers so to adjustthe HLB value to fall within these ranges, thus to achieve an equaleffect.

Silicone emulsifiers are preferred. A wide variety of siliconeemulsifiers are useful herein. These silicone emulsifiers are typicallyorganically modified organopolysiloxanes, also known to those skilled inthe art as silicone surfactants. Useful silicone emulsifiers includedimethicone copolyols. These materials are polydimethyl siloxanes whichhave been modified to include polyether side chains such as polyethyleneoxide chains, polypropylene oxide chains, mixtures of these chains, andpolyether chains containing moieties derived from both ethylene oxideand propylene oxide. Other examples include alkyl-modified dimethiconecopolyols, i.e., compounds which contain C₂-C₃₀ pendant side chains.Still other useful dimethicone copolyols include materials havingvarious cationic, anionic, amphoteric, and zwitterionic pendantmoieties.

Also useful herein, although not strictly classified as dimethiconecopolyols, are silicone surfactants.

Nonlimiting examples of dimethicone copolyols and other siliconesurfactants useful as emulsifiers herein include polydimethylsiloxanepolyether copolymers with pendant polyethylene oxide sidechains,polydimethylsiloxane polyether copolymers with pendant polypropyleneoxide sidechains, polydimethylsiloxane polyether copolymers with pendantmixed polyethylene oxide and polypropylene oxide sidechains,polydimethylsiloxane polyether copolymers with pendant mixedpoly(ethylene)(propylene)oxide sidechains, polydimethylsiloxanepolyether copolymers with pendant organobetaine sidechains,polydimethylsiloxane polyether copolymers with pendant carboxylatesidechains, polydimethylsiloxane polyether copolymers with pendantquaternary ammonium sidechains; and also further modifications of thepreceding copolymers containing pendant C₂-C₃₀ straight, branched, orcyclic alkyl moieties. Examples of commercially available dimethiconecopolyols useful herein sold by Dow Corning Corporation are Dow Corning®190, 193, Q2-5220, 2501 Wax, 2-5324 fluid, and 3225C (this latermaterial being sold as a mixture with cyclomethicone). Cetyl dimethiconecopolyol is commercially available as a mixture with polyglyceryl-4isostearate (and) hexyl laurate and is sold under the tradename ABIL®WE-09 (available from Goldschmidt). Cetyl dimethicone copolyol is alsocommercially available as a mixture with hexyl laurate (and)polyglyceryl-3 oleate (and) cetyl dimethicone and is sold under thetradename ABIL® WS-08 (also available from Goldschmidt). Othernonlimiting examples of dimethicone copolyols also include lauryldimethicone copolyol, dimethicone copolyol acetate, diemethiconecopolyol adipate, dimethicone copolyolamine, dimethicone copolyolbehenate, dimethicone copolyol butyl ether, dimethicone copolyol hydroxystearate, dimethicone copolyol isostearate, dimethicone copolyollaurate, dimethicone copolyol methyl ether, dimethicone copolyolphosphate, and dimethicone copolyol stearate. See International CosmeticIngredient Dictionary, Fifth Edition, 1993.

Dimethicone copolyol emulsifiers useful herein are described, forexample, in U.S. Pat. No. 4,960,764, to Figueroa, Jr. et al., issuedOct. 2, 1990; European Patent No. EP 330,369, to SanoGueira, publishedAug. 30, 1989; G. H. Dahms, et al., “New Formulation PossibilitiesOffered by Silicone Copolyols,” Cosmetics & Toiletries, vol. 110, pp.91-100, March 1995; M. E. Carlotti et al., “Optimization of W/O-SEmulsions And Study Of The Quantitative Relationships Between EsterStructure And Emulsion Properties,” J. Dispersion Science AndTechnology, 13(3), 315-336 (1992); P. Hameyer, “ComparativeTechnological Investigations of Organic and Organosilicone Emulsifiersin Cosmetic Water-in-Oil Emulsion Preparations,” HAPPI 28(4), pp. 88-128(1991); J. Smid-Korbar et al., “Efficiency and usability of siliconesurfactants in emulsions,” Provisional Communication, InternationalJournal of Cosmetic Science, 12, 135-139 (1990); and D. G. Krzysik etal., “A New Silicone Emulsifier For Water-in-Oil Systems,” Drug andCosmetic Industry, vol. 146(4) pp. 28-81 (April 1990).

Among the non-silicone-containing emulsifiers useful herein are variousnon-ionic and anionic emulsifying agents such as sugar esters andpolyesters, alkoxylated sugar esters and polyesters, C₁-C₃₀ fatty acidesters of C₁-C₃₀ fatty alcohols, alkoxylated derivatives of C₁-C₃₀ fattyacid esters of C₁-C₃₀ fatty alcohols, alkoxylated ethers of C₁-C₃₀ fattyalcohols, polyglyceryl esters of C₁-C₃₀ fatty acids, C₁-C₃₀ esters ofpolyols, C₁-C₃₀ ethers of polyols, alkyl phosphates, polyoxyalkylenefatty ether phosphates, fatty acid amides, acyl lactylates, soaps, andmixtures thereof. Other suitable emulsifiers are described, for example,in McCutcheon's, Detergents and Emulsifiers, North American Edition(1986), published by Allured Publishing Corporation; U.S. Pat. No.5,011,681 to Ciotti et al., issued Apr. 30, 1991; U.S. Pat. No.4,421,769 to Dixon et al., issued Dec. 20, 1983; and U.S. Pat. No.3,755,560 to Dickert et al., issued Aug. 28, 1973.

Nonlimiting examples of these non-silicon-containing emulsifiersinclude: polyethylene glycol 20 sorbitan monolaurate (Polysorbate 20),polyethylene glycol 5 soya sterol, Steareth-20, Ceteareth-20, PPG-2methyl glucose ether distearate, Ceteth-10, Polysorbate 80, cetylphosphate, potassium cetyl phosphate, diethanolamine cetyl phosphate,Polysorbate 60, glyceryl stearate, PEG-100 stearate, polyoxyethylene 20sorbitan trioleate (Polysorbate 85), sorbitan monolaurate,polyoxyethylene 4 lauryl ether sodium stearate, polyglyceryl-4isostearate, hexyl laurate, steareth-20, ceteareth-20, PPG-2 methylglucose ether distearate, ceteth-10, diethanolamine cetyl phosphate,glyceryl stearate, PEG-100 stearate, and mixtures thereof.

Oil-in-Water Emulsions

Other preferred carriers include oil-in-water emulsions, having acontinuous aqueous phase and a water-insoluble phase (“oil phase”)dispersed therein. Examples of suitable oil-in-water emulsion aredescribed in U.S. Pat. No. 5,073,371, to Turner, D. J. et al., issuedDec. 17, 1991, and U.S. Pat. No. 5,073,372, to Turner, D. J. et al.,issued Dec. 17, 1991. A preferred oil-in-water emulsion, containing astructuring agent, hydrophilic surfactant and water, is described indetail hereinafter.

(1) Structuring Agent

A preferred oil-in-water emulsion contains a structuring agent to assistin the formation of a liquid crystalline gel network structure. Withoutbeing limited by theory, it is believed that the structuring agentassists in providing rheological characteristics to the compositionwhich contribute to the stability of the composition. The structuringagent may also function as an emulsifier or surfactant. The emulsionaccording to the invention contains from about 0.5% to about 20%, morepreferably from about 1% to about 10%, even more preferably from about1% to about 5%, by weight of the composition, of a structuring agent.

The preferred structuring agents of the present invention includestearic acid, palmitic acid, stearyl alcohol, cetyl alcohol, behenylalcohol, stearic acid, palmitic acid, the polyethylene glycol ether ofstearyl alcohol having an average of about 1 to about 21 ethylene oxideunits, the polyethylene glycol ether of cetyl alcohol having an averageof about 1 to about 7 ethylene oxide units, and mixtures thereof. Otherstructuring agents used in the present invention are selected fromstearyl alcohol, cetyl alcohol, behenyl alcohol, the polyethylene glycolether of stearyl alcohol having an average of about 2 ethylene oxideunits (steareth-2), the polyethylene glycol ether of stearyl alcoholhaving an average of about 21 ethylene oxide units (steareth-21), andmixtures thereof. The structuring agents can also be one which isselected from stearic acid, palmitic acid, stearyl alcohol, cetylalcohol, behenyl alcohol, steareth-2, steareth-21, and mixtures thereof.

(2) Hydrophilic Surfactant

The preferred oil-in-water emulsions contain from about 0.05% to about10%, preferably from about 1% to about 6%, and more preferably fromabout 1% to about 5% (percentages by weight of the total emulsion) of atleast one hydrophilic surfactant which can disperse the hydrophobicmaterials in the water phase. The surfactant, at a minimum, must behydrophilic enough to disperse in water.

Preferred hydrophilic surfactants are selected from nonionicsurfactants. Among the nonionic surfactants that are useful herein arethose that can be broadly defined as condensation products of long chainalcohols, e.g. C₈₋₃₀ alcohols, with sugar or starch polymers, i.e.,glycosides. These compounds can be represented by the formula (S)_(n)—O—R wherein S is a sugar moiety such as glucose, fructose, mannose, andgalactose; n is an integer of from about 1 to about 1000, and R is aC₈₋₃₀ alkyl group. Examples of long chain alcohols from which the alkylgroup can be derived include decyl alcohol, cetyl alcohol, stearylalcohol, lauryl alcohol, myristyl alcohol, oleyl alcohol, and the like.Preferred examples of these surfactants include those wherein S is aglucose moiety, R is a C₈₋₂₀ alkyl group, and n is an integer of fromabout 1 to about 9. Commercially available examples of these surfactantsinclude decyl polyglucoside (available as APG 325 CS from Henkel) andlauryl polyglucoside (available as APG 600 CS and 625 CS from Henkel).

Other useful nonionic surfactants include the condensation products ofalkylene oxides with fatty acids (i.e. alkylene oxide esters of fattyacids). These materials have the general formula RCO(X)_(n)OH wherein Ris a C₁₀₋₃₀ alkyl group, X is —OCH₂CH₂— (i.e. derived from ethyleneglycol or oxide) or —OCH₂CHCH₃— (i.e. derived from propylene glycol oroxide), and n is an integer from about 6 to about 200. Other nonionicsurfactants are the condensation products of alkylene oxides with 2moles of fatty acids (i.e. alkylene oxide diesters of fatty acids).These materials have the general formula RCO(X)_(n)OOCR wherein R is aC₁₀₋₃₀ alkyl group, X is —OCH₂CH₂— (i.e. derived from ethylene glycol oroxide) or —OCH₂CHCH₃— (i.e. derived from propylene glycol or oxide), andn is an integer from about 6 to about 100. Other nonionic surfactantsare the condensation products of alkylene oxides with fatty alcohols(i.e. alkylene oxide ethers of fatty alcohols). These materials have thegeneral formula R(X).sub.n OR′ wherein R is a C10-30 alkyl group, X is—OCH₂CH₂— (i.e. derived from ethylene glycol or oxide) or —OCH₂CHCH₃—(i.e. derived from propylene glycol or oxide), and n is an integer fromabout 6 to about 100 and R′ is H or a C₁₀₋₃₀ alkyl group. Still othernonionic surfactants are the condensation products of alkylene oxideswith both fatty acids and fatty alcohols [i.e. wherein the polyalkyleneoxide portion is esterified on one end with a fatty acid and etherified(i.e. connected via an ether linkage) on the other end with a fattyalcohol]. These materials have the general formula RCO(X)_(n)OR′ whereinR and R′ are C₁₀₋₃₀ alkyl groups, X is —OCH₂ CH₂ (i.e. derived fromethylene glycol or oxide) or —OCH₂CHCH₃— (derived from propylene glycolor oxide), and n is an integer from about 6 to about 100. Nonlimitingexamples of these alkylene oxide derived nonionic surfactants includeceteth-6, ceteth-10, ceteth-12, ceteareth-6, ceteareth-10, ceteareth-12,steareth-6, steareth-10, steareth-12, steareth-21, PEG-6 stearate,PEG-10 stearate, PEG-100 stearate, PEG-12 stearate, PEG-20 glycerylstearate, PEG-80 glyceryl tallowate, PEG-10 glyceryl stearate, PEG-30glyceryl cocoate, PEG-80 glyceryl cocoate, PEG-200 glyceryl tallowate,PEG-8 dilaurate, PEG-10 distearate, and mixtures thereof.

Preferred among the nonionic surfactants are those selected from thegroup consisting of steareth-21, ceteareth-20, ceteareth-12, sucrosecocoate, steareth-100, PEG-100 stearate, and mixtures thereof.

Other nonionic surfactants suitable for use herein include sugar estersand polyesters, alkoxylated sugar esters and polyesters, C₁-C₃₀ fattyacid esters of C₁-C₃₀ fatty alcohols, alkoxylated derivatives of C₁-C₃₀fatty acid esters of C₁-C₃₀ fatty alcohols, alkoxylated ethers of C₁-C₃₀fatty alcohols, polyglyceryl esters of C₁-C₃₀ fatty acids, C₁-C₃₀ estersof polyols, C₁-C₃₀ ethers of polyols, alkyl phosphates, polyoxyalkylenefatty ether phosphates, fatty acid amides, acyl lactylates, and mixturesthereof. Nonlimiting examples of these emulsifiers include: polyethyleneglycol 20 sorbitan monolaurate (Polysorbate 20), polyethylene glycol 5soya sterol, Steareth-20, Ceteareth-20, PPG-2 methyl glucose etherdistearate, Ceteth-10, Polysorbate 80, cetyl phosphate, potassium cetylphosphate, diethanolamine cetyl phosphate, Polysorbate 60, glycerylstearate, polyoxyethylene 20 sorbitan trioleate (Polysorbate 85),sorbitan monolaurate, polyoxyethylene 4 lauryl ether sodium stearate,polyglyceryl-4 isostearate, hexyl laurate, PPG-2 methyl glucose etherdistearate, PEG-100 stearate, and mixtures thereof.

Another emulsifier useful herein are fatty acid ester blends based on amixture of sorbitan or sorbitol fatty acid ester and sucrose fatty acidester, the fatty acid in each instance being preferably C₈-C₂₄, morepreferably C₁₀-C₂₀. The preferred fatty acid ester emulsifier is a blendof sorbitan or sorbitol C₁₆-C₂₀ fatty acid ester with sucrose C₁₀-C₁₆fatty acid ester, especially sorbitan stearate and sucrose cocoate. Thisis commercially available from ICI under the trade name Arlatone 2121.

Other suitable surfactants useful herein include a wide variety ofcationic, anionic, zwitterionic, and amphoteric surfactants such as areknown in the art. See, e.g., McCutcheon's, Detergents and Emulsifiers,North American Edition (1986), published by Allured PublishingCorporation; U.S. Pat. No. 5,011,681 to Ciotti et al., issued Apr. 30,1991; U.S. Pat. No. 4,421,769 to Dixon et al., issued Dec. 20, 1983; andU.S. Pat. No. 3,755,560 to Dickert et al., issued Aug. 28, 1973; thesefour references are incorporated herein by reference in their entirety.The hydrophilic surfactants useful herein can contain a singlesurfactant, or any combination of suitable surfactants. The exactsurfactant (or surfactants) chosen will depend upon the pH of thecomposition and the other components present.

Also useful herein are cationic surfactants, especially dialkylquaternary ammonium compounds, examples of which are described in U.S.Pat. Nos. 5,151,209; 5,151,210; 5,120,532; 4,387,090; 3,155,591;3,929,678; 3,959,461; McCutcheon's, Detergents & Emulsifiers, (NorthAmerican edition 1979) M.C. Publishing Co.; and Schwartz, et al.,Surface Active Agents, Their Chemistry and Technology, New York:Interscience Publishers, 1949; which descriptions are incorporatedherein by reference.

Alternatively, other useful cationic emulsifiers include amino-amides.Nonlimiting examples of these cationic emulsifiers includestearamidopropyl PG-dimonium chloride phosphate, behenamidopropyl PGdimonium chloride, stearamidopropyl ethyldimonium ethosulfate,stearamidopropyl dimethyl(myristyl acetate)ammonium chloride,stearamidopropyl dimethyl cetearyl ammonium tosylate, stearamidopropyldimethyl ammonium chloride, stearamidopropyl dimethyl ammonium lactate,and mixtures thereof. Especially preferred is behenamidopropyl PGdimonium chloride.

Nonlimiting examples of quaternary ammonium salt cationic surfactantsinclude those selected from cetyl ammonium chloride, cetyl ammoniumbromide, lauryl ammonium chloride, lauryl ammonium bromide, stearylammonium chloride, stearyl ammonium bromide, cetyl dimethyl ammoniumchloride, cetyl dimethyl ammonium bromide, lauryl dimethyl ammoniumchloride, lauryl dimethyl ammonium bromide, stearyl dimethyl ammoniumchloride, stearyl dimethyl ammonium bromide, cetyl trimethyl ammoniumchloride, cetyl trimethyl ammonium bromide, lauryl trimethyl ammoniumchloride, lauryl trimethyl ammonium bromide, stearyl trimethyl ammoniumchloride, stearyl trimethyl ammonium bromide, lauryl dimethyl ammoniumchloride, stearyl dimethyl cetyl ditallow dimethyl ammonium chloride,dicetyl ammonium chloride, dicetyl ammonium bromide, dilauryl ammoniumchloride, dilauryl ammonium bromide, distearyl ammonium chloride,distearyl ammonium bromide, dicetyl methyl ammonium chloride, dicetylmethyl ammonium bromide, dilauryl methyl ammonium chloride, dilaurylmethyl ammonium bromide, distearyl methyl ammonium chloride, distearylmethyl ammonium bromide, and mixtures thereof. Additional quaternaryammonium salts include those wherein the C₁₂ to C₃₀ alkyl carbon chainis derived from a tallow fatty acid or from a coconut fatty acid. Theterm “tallow” refers to an alkyl group derived from tallow fatty acids(usually hydrogenated tallow fatty acids), which generally have mixturesof alkyl chains in the C₁₆ to C₁₈ range. The term “coconut” refers to analkyl group derived from a coconut fatty acid, which generally havemixtures of alkyl chains in the C₁₂ to C₁₄ range. Examples of quaternaryammonium salts derived from these tallow and coconut sources includeditallow dimethyl ammonium chloride, ditallow dimethyl ammonium methylsulfate, di(hydrogenated tallow)dimethyl ammonium chloride,di(hydrogenated tallow)dimethyl ammonium acetate, ditallow dipropylammonium phosphate, ditallow dimethyl ammonium nitrate,di(coconutalkyl)dimethyl ammonium chloride, di(coconutalkyl)dimethylammonium bromide, tallow ammonium chloride, coconut ammonium chloride,stearamidopropyl PG-dimonium chloride phosphate, stearamidopropylethyldimonium ethosulfate, stearamidopropyl dimethyl(myristylacetate)ammonium chloride, stearamidopropyl dimethyl cetearyl ammoniumtosylate, stearamidopropyl dimethyl ammonium chloride, stearamidopropyldimethyl ammonium lactate, and mixtures thereof. An example of aquaternary ammonium compound having an alkyl group with an ester linkageis ditallowyl oxyethyl dimethyl ammonium chloride.

More preferred cationic surfactants are those selected frombehenamidopropyl PG dimonium chloride, dilauryl dimethyl ammoniumchloride, distearyl dimethyl ammonium chloride, dimyristyl dimethylammonium chloride, dipalmityl dimethyl ammonium chloride, distearyldimethyl ammonium chloride, stearamidopropyl PG-dimonium chloridephosphate, stearamidopropyl ethyldiammonium ethosulfate,stearamidopropyl dimethyl(myristyl acetate)ammonium chloride,stearamidopropyl dimethyl cetearyl ammonium tosylate, stearamidopropyldimethyl ammonium chloride, stearamidopropyl dimethyl ammonium lactate,and mixtures thereof.

Still more preferred cationic surfactants are those selected frombehenamidopropyl PG dimonium chloride, dilauryl dimethyl ammoniumchloride, distearyl dimethyl ammonium chloride, dimyristyl dimethylammonium chloride, dipalmityl dimethyl ammonium chloride, and mixturesthereof.

A preferred combination of cationic surfactant and structuring agent isbehenamidopropyl PG dimonium chloride and/or behenyl alcohol, whereinthe ratio is preferably optimized to maintain to enhance physical andchemical stability, especially when such a combination contains ionicand/or highly polar solvents. This combination is especially useful fordelivery of sunscreening agents such as zinc oxide and octylmethoxycinnamate.

A wide variety of anionic surfactants are also useful herein. See, e.g.,U.S. Pat. No. 3,929,678, to Laughlin et al., issued Dec. 30, 1975, whichis incorporated herein by reference in its entirety. Nonlimitingexamples of anionic surfactants include the alkoyl isethionates, and thealkyl and alkyl ether sulfates. The alkoyl isethionates typically havethe formula RCO—OCH₂CH₂SO₃M wherein R is alkyl or alkenyl of from about10 to about 30 carbon atoms, and M is a water-soluble cation such asammonium, sodium, potassium and triethanolamine. Nonlimiting examples ofthese isethionates include those alkoyl isethionates selected fromammonium cocoyl isethionate, sodium cocoyl isethionate, sodium lauroylisethionate, sodium stearoyl isethionate, and mixtures thereof.

The alkyl and alkyl ether sulfates typically have the respectiveformulae ROSO₃M and RO(C₂H₄O)_(x)SO₃M, wherein R is alkyl or alkenyl offrom about 10 to about 30 carbon atoms, x is an integer of from about 1to about 10, and M is a water-soluble cation such as ammonium, sodium,potassium and triethanolamine.

Other anionic materials useful herein are soaps (i.e. alkali metalsalts, e.g., sodium or potassium salts) of fatty acids, typically havingfrom about 8 to about 24 carbon atoms, preferably from about 10 to about20 carbon atoms. The fatty acids used in making the soaps can beobtained from natural sources such as, for instance, plant oranimal-derived glycerides (e.g., palm oil, coconut oil, soybean oil,castor oil, tallow, lard, etc.) The fatty acids can also besynthetically prepared. Soaps are described in more detail in U.S. Pat.No. 4,557,853.

Amphoteric and zwitterionic surfactants are also useful herein. Examplesof amphoteric and zwitterionic surfactants which can be used in thecosmetic compositions of the present invention are those which arebroadly described as derivatives of aliphatic secondary and tertiaryamines in which the aliphatic radical can be straight or branched chainand wherein one of the aliphatic substituents contains from about 8 toabout 22 carbon atoms (preferably C₈-C₁₈) and one contains an anionicwater solubilizing group, e.g., carboxy, sulfonate, sulfate, phosphate,or phosphonate. Examples are alkyl imino acetates, and iminodialkanoatesand aminoalkanoates of the formulas RN[CH₂)_(m)CO₂M]₂ andRNH(CH₂)_(m)CO₂M wherein m is an integer of from 1 to 4, R is a C₈-C₂₂alkyl or alkenyl, and M is H, alkali metal, alkaline earth metalammonium, or alkanolammonium. Also included are imidazolinium andammonium derivatives. Specific examples of suitable amphotericsurfactants include sodium 3-dodecyl-aminopropionate, sodium3-dodecylaminopropane sulfonate, N-alkyltaurines such as the oneprepared by reacting dodecylamine with sodium isethionate according tothe teaching of U.S. Pat. No. 2,658,072 which is incorporated herein byreference in its entirety; N-higher alkyl aspartic acids such as thoseproduced according to the teaching of U.S. Pat. No. 2,438,091 which isincorporated herein by reference in its entirety; and the products soldunder the trade name “Miranol” and described in U.S. Pat. No. 2,528,378,which is incorporated herein by reference in its entirety. Otherexamples of useful amphoterics include phosphates, such as coamidopropylPG-dimonium chloride phosphate (commercially available as Monaquat PTC,from Mona Corp.).

Other amphoteric or zwitterionic surfactants useful herein includebetaines. Examples of betaines include the higher alkyl betaines, suchas coco dimethyl carboxymethyl betaine, lauryl dimethyl carboxymethylbetaine, lauryl dimethyl alphacarboxyethyl betaine, cetyl dimethylcarboxymethyl betaine, cetyl dimethyl betaine (available as Lonzaine16SP from Lonza Corp.), lauryl bis-(2-hydroxyethyl)carboxymethylbetaine, stearyl bis-(2-hydroxypropyl)carboxy-methyl betaine, oleyldimethyl gamma-carboxypropyl betaine, laurylbis-(2-hydroxypropyl)-alpha-carboxyethyl betaine, coco dimethylsulfopropyl betaine, stearyl dimethyl sulfopropyl betaine, lauryldimethyl sulfoethyl betaine, lauryl bis-(2-hydroxyethyl)sulfopropylbetaine, and amidobetaines and amidosulfobetaines (wherein theRCONH(CH₂)₃ radical is attached to the nitrogen atom of the betaine),oleyl betaine (available as amphoteric Velvetex OLB-50 from Henkel), andcocamidopropyl betaine (available as Velvetex BK-35 and BA-35 fromHenkel).

Other useful amphoteric and zwitterionic surfactants include thesultaines and hydroxysultaines such as cocamidopropyl hydroxysultaine(available as Mirataine CBS from Rhone-Poulenc), and the alkanoylsarcosinates corresponding to the formula RCON(CH₃)CH₂CH₂CO₂M wherein Ris alkyl or alkenyl of about 10 to about 20 carbon atoms, and M is awater-soluble cation such as ammonium, sodium, potassium andtrialkanolamine (e.g., triethanolamine), a preferred example of which issodium lauroyl sarcosinate.

(3) Water

The preferred oil-in-water emulsion contains from about 10% to about98%, preferably from about 20% to about 95%, more preferably from about30% to about 90% water by weight of the liquid carrier.

The hydrophobic phase is dispersed in the continuous aqueous phase. Thehydrophobic phase may contain water insoluble or partially solublematerials such as are known in the art, including but not limited to thesilicones described herein in reference to silicone-in-water emulsions,and other oils and lipids such as described above in reference toemulsions.

The cosmetic compositions of the present invention, including but notlimited to lotions and creams, may contain a dermatologically acceptableemollient. Such compositions preferably contain from about 1% to about50% of the emollient. As used herein, “emollient” refers to a materialuseful for the prevention or relief of dryness, as well as for theprotection of the skin. A wide variety of suitable emollients are knownand may be used herein. Sagarin, Cosmetics, Science and Technology, 2ndEdition, Vol. 1, pp. 32-43 (1972), incorporated herein by reference,contains numerous examples of materials suitable as an emollient. Apreferred emollient is glycerin. Glycerin is preferably used in anamount of from about 0.001 to about 30%, more preferably from about 0.01to about 20%, still more preferably from about 0.1 to about 10%, e.g.,5%.

Lotions and creams according to the present invention generally containa solution carrier system and one or more emollients. Lotions and creamstypically contain from about 1% to about 50%, preferably from about 1%to about 20%, of emollient; from about 50% to about 90%, preferably fromabout 60% to about 80%, water; and the vitamin B3 compound and skin careactive substance in the above described amounts. Creams are generallythicker than lotions.

Ointments of the present invention may contain animal or vegetable oilsor semi-solid hydrocarbons (oleaginous); absorption ointment bases whichabsorb water to form emulsions; or water soluble carriers, e.g., a watersoluble solution carrier. Ointments may further contain a thickeningagent, such as described in Sagarin, Cosmetics, Science and Technology,2nd Edition, Vol. 1, pp. 72-73 (1972), incorporated herein by reference,and/or an emollient. For example, an ointment may contain from about 2%to about 10% of an emollient; from about 0.1% to about 2% of athickening agent; and the vitamin B3 compound and skin care activesubstance in the above described amounts.

Foundation carriers according to the present invention may contain apowder system selected from talc or mica, colorants, and the plantextract, with or without additional solvents. As used herein, the term“foundation” refers to a liquid, semi-liquid, semi-solid, or solid skincosmetic which includes, but is not limited to lotions, creams, gels,pastes, cakes, and the like. Typically the foundation is used over alarge area of the skin, such as the face, to provide a particulareffect. Foundations are typically used to provide an adherent base forcolor cosmetics such as rouge, blusher, powder and the like, and tend tohide skin imperfections and impart a smooth, even appearance to theskin. Foundations in the present invention include a dermatologicallyacceptable carrier and may include conventional ingredients such asoils, colorants, pigments, emollients, fragrances, waxes, stabilizers,and the like. Exemplary carriers and such other ingredients which aresuitable for use herein are described, for example, in PCT Application,WO 96/33689, to Canter, et al., published on Oct. 31, 1996 and U.K.Patent, GB 2274585, issued on Aug. 3, 1994.

The cosmetic composition of the invention may contain one or moreadditional skin care actives.

In a preferred embodiment, where the composition is to be in contactwith human keratinous tissue, the additional components should besuitable for application to keratinous tissue, that is, whenincorporated into the composition they are suitable for use in contactwith human keratinous tissue without undue toxicity, incompatibility,instability, allergic response, and the like within the scope of soundmedical judgment. The CTFA Cosmetic Ingredient Handbook, Second Edition(1992) describes a wide variety of nonlimiting cosmetic andpharmaceutical ingredients commonly used in the skin care industry,which are suitable for use in the compositions of the present invention.Examples of these ingredient classes include: abrasives, absorbents,aesthetic components such as fragrances, pigments, colorings/colorants,essential oils, skin sensates, astringents, etc. (e.g., clove oil,menthol, camphor, eucalyptus oil, eugenol, menthyl lactate, witch hazeldistillate), anti-acne agents, anti-caking agents, antifoaming agents,antimicrobial agents (e.g., iodopropyl butylcarbamate), antioxidants,binders, biological additives, buffering agents, bulking agents,chelating agents, chemical additives, colorants, cosmetic astringents,cosmetic biocides, denaturants, drug astringents, external analgesics,film formers or materials, e.g., polymers, for aiding the film-formingproperties and substantivity of the composition (e.g., copolymer ofeicosene and vinyl pyrrolidone), opacifying agents, pH adjusters,propellants, reducing agents, sequestrants, skin bleaching andlightening agents (e.g., hydroquinone, kojic acid, ascorbic acid,magnesium ascorbyl phosphate, ascorbyl glucosamine, ascorbyl glucoside),skin-conditioning agents (e.g., humectants, including miscellaneous andocclusive), skin soothing and/or healing agents (e.g., panthenol andderivatives (e.g., ethyl panthenol), aloe vera, pantothenic acid and itsderivatives, allantoin, bisabolol, and dipotassium glycyrrhizinate),skin treating agents, thickeners, and vitamins and derivatives thereof.

Desquamation Actives

A safe and effective amount of a desquamation active may be added to thecompositions of the present invention, more preferably from about 0.1%to about 10%, even more preferably from about 0.2% to about 5%, alsopreferably from about 0.5% to about 4%, by weight of the composition.Desquamation actives used in the present invention are favorable to theskin appearance. For example, the desquamation actives tend to improvethe texture of the skin (e.g., smoothness). One desquamation system thatis suitable for use herein contains sulfhydryl compounds andzwitterionic surfactants and is described in U.S. Pat. No. 5,681,852, toBissett, incorporated herein by reference. Another desquamation systemthat is suitable for use herein contains salicylic acid and zwitterionicsurfactants and is described in U.S. Pat. No. 5,652,228 to Bissett,incorporated herein by reference. Zwitterionic surfactants such as thosedescribed in these applications are also useful as desquamatory agentsherein, with cetyl betaine being particularly preferred.

Anti-Acne Actives

The compositions of the present invention may contain a safe andeffective amount of one or more anti-acne actives. Examples of usefulanti-acne actives include resorcinol, sulfur, salicylic acid, benzoylperoxide, erythromycin, zinc, etc., which are described in furtherdetail in U.S. Pat. No. 5,607,980, issued to McAtee et al, on Mar. 4,1997.

Anti-Wrinkle Actives/Anti-Atrophy Actives

The compositions of the present invention may further contain a safe andeffective amount of one or more anti-wrinkle actives or anti-atrophyactives. Exemplary anti-wrinkle/anti-atrophy actives suitable for use inthe compositions of the present invention include sulfur-containing Dand L amino acids and their derivatives and salts, particularly theN-acetyl derivatives, a preferred example of which isN-acetyl-L-cysteine; thiols, e.g. ethane thiol; hydroxy acids (e.g.,alpha-hydroxy acids such as lactic acid and glycolic acid orbeta-hydroxy acids such as salicylic acid and salicylic acid derivativessuch as the octanoyl derivative), phytic acid, lipoic acid;lysophosphatidic acid, skin peel agents (e.g., phenol and the like),vitamin B₃ compounds and retinoids which can improve the keratinoustissue appearance, especially in regulating keratinous tissue condition.

a) Vitamin B₃ Compounds

The compositions of the present invention may contain a safe andeffective amount of a vitamin B₃ compound. Vitamin B₃ compounds areuseful for regulating skin condition as described in co-pending U.S.application Ser. No. 08/834,010, filed Apr. 11, 1997 (corresponding tointernational publication WO 97/39733 A1, published Oct. 30, 1997). Whenvitamin B₃ compounds are present in the compositions of the instantinvention, the compositions preferably contain from about 0.01% to about50%, more preferably from about 0.1% to about 10%, even more preferablyfrom about 0.5% to about 10%, and still more preferably from about 1% toabout 5%, still more preferably from about 2% to about 5%, by weight ofthe composition, of the vitamin B₃ compound.

Exemplary derivatives of the foregoing vitamin B₃ compounds includenicotinic acid esters, including non-vasodilating esters of nicotinicacid (e.g., tocopheryl nicotinate), nicotinyl amino acids, nicotinylalcohol esters of carboxylic acids, nicotinic acid N-oxide andniacinamide N-oxide.

Examples of suitable vitamin B₃ compounds are well known in the art andare commercially available from a number of sources, e.g., the SigmaChemical Company (St. Louis, Mo.); ICN Biomedicals, Inc. (Irvin, Calif.)and Aldrich Chemical Company (Milwaukee, Wis.).

The vitamin compounds may be included as the substantially purematerial, or as an extract obtained by suitable physical and/or chemicalisolation from natural (e.g., plant) sources.

b) Retinoids

The compositions of the present invention may also contain a retinoidcomposition. As used herein, “retinoid” includes all natural and/orsynthetic analogs of Vitamin A or retinol-like compounds which possessthe biological activity of Vitamin A in the skin as well as thegeometric isomers and stereoisomers of these compounds. The retinoid ispreferably retinol, retinol esters (e.g., C₂-C₂₂ alkyl esters ofretinol, including retinyl palmitate, retinyl acetate, retinylpropionate), retinal, and/or retinoic acid (including all-trans retinoicacid and/or 1,3-cis-retinoic acid), more preferably retinoids other thanretinoic acid. These compounds are well known in the art and arecommercially available from a number of sources, e.g., Sigma ChemicalCompany (St. Louis, Mo.), and Boerhinger Mannheim (Indianapolis, Ind.).Other retinoids which are useful herein are described in U.S. Pat. No.4,677,120, issued Jun. 30, 1987 to Parish et al.; U.S. Pat. No.4,885,311, issued Dec. 5, 1989 to Parish et al.; U.S. Pat. No.5,049,584, issued Sep. 17, 1991 to Purcell et al.; U.S. Pat. No.5,124,356, issued Jun. 23, 1992 to Purcell et al.; and U.S. Pat. No.Reissue 34,075, issued Sep. 22, 1992 to Purcell et al. Other suitableretinoids are tocopheryl-retinoate [tocopherol ester of retinoic acid(trans- or cis-), adapalene{6-[3-(1-adamantyl)-4-methoxyphenyl]-2-naphthoic acid}, and tazarotene(ethyl 6-[2-(4,4-dimethylthiochroman-6-yl)-ethynyl]nicotinate).Preferred retinoids are retinol, retinyl palmitate, retinyl acetate,retinyl propionate, retinal and combinations thereof.

The retinoid may be included as the substantially pure material, or asan extract obtained by suitable physical and/or chemical isolation fromnatural (e.g., plant) sources. The retinoid is preferably substantiallypure, more preferably essentially pure.

The compositions of this invention may contain a safe and effectiveamount of the retinoid, such that the resultant composition is safe andeffective for regulating keratinous tissue condition, preferably forregulating appearance and/or tactile feeling in skin, more preferablyfor regulating signs of skin aging. The compositions preferably containfrom about 0.005% to about 2%, more preferably 0.01% to about 2%,retinoid. Retinol is preferably used in an amount of from or about 0.01%to or about 0.15%; retinol esters are preferably used in an amount offrom about 0.01% to about 2% (e.g., about 1%); retinoic acids arepreferably used in an amount of from about 0.01% to about 0.25%;tocopheryl-retinoate, adapalene, and tazarotene are preferably used inan amount of from about 0.01% to about 2%.

Where the compositions of the present invention contain both a retinoidand a Vitamin B₃ compound, the retinoid is preferably used in the aboveamounts, and the vitamin B₃ compound is preferably used in an amount offrom about 0.1% to about 10%, more preferably from about 2% to about 5%.

c) Hydroxy Acids

The compositions of the present invention may contain a safe andeffective amount of a Hydroxy Acid. Preferred hydroxy acids for use inthe compositions of the present invention include salicylic acid andsalicylic acid derivatives. When present in the compositions of thepresent invention, salicylic acid is preferably used in an amount offrom about 0.01% to about 50%, more preferably from about 0.1% to about20%, even more preferably from about 0.1% to about 10%, still morepreferably from about 0.5% to about 5%, and still more preferably fromabout 0.5% to about 2%.

Peptides

Peptides, including but not limited to, di-, tri-, tetra-, andpentapeptides and derivatives thereof, may be included in thecompositions of the present invention in amounts that are safe andeffective. As used herein, “peptides” refers to both the naturallyoccurring peptides and synthesized peptides. Also useful herein arenaturally occurring and commercially available compositions that containpeptides.

Suitable dipeptides for use herein include Carnosine (beta-ala-his).Suitable tripeptides for use herein include, gly-his-lys, arg-lys-arg,his-gly-gly. Preferred tripeptides and derivatives thereof includepalmitoyl-gly-his-lys, which may be purchased as Biopeptide CL® (100 ppmof palmitoyl-gly-his-lys commercially available from Sederma, France);Peptide CK (arg-lys-arg); Peptide CK+ (ac-arg-lys-arg-NH₂); and a copperderivative of his-gly-gly sold commercially as Iamin, from Sigma (St.Louis, Mo.). Suitable tetrapeptides for use herein include Peptide E,arg-ser-arg-lys (SEQ ID NO:1). Suitable pentapeptides for use hereininclude lys-thr-thr-lys-ser. A preferred commercially availablepentapeptide derivative composition is Matrixyl®, which contains 100 ppmpalmitoyl-lys-thr-thr-lys-ser (SEQ ID NO:2, commercially available fromSederma France).

Preferably, the peptide is selected from palmitoyl-lys-thr-thr-lys-ser,palmitoyl-gly-his-lys, beta-ala-his, their derivatives, and combinationsthereof. More preferably, the peptide is selected frompalmitoyl-lys-thr-thr-lys-ser, palmitoyl-gly-his-lys, their derivatives,and combinations thereof. Even more preferably, the peptide is selectedfrom palmitoyl-lys-thr-thr-lys-ser and derivatives thereof.

When included in the present compositions, peptides are preferablyincluded in amounts of from about 1×10⁻⁶% to about 10%, more preferablyfrom about 1×10⁻⁶% to about 0.1%, even more preferably from about1×10⁻⁵% to about 0.01%, by weight of the composition. In certaincompositions where the peptide is Carnosine®, the compositionspreferably contain from about 0.1% to about 5%, by weight of thecomposition, of such peptides. In other embodiments wherein thepeptide-containing compositions, Matrixyl®, and/or Biopeptide CL® areincluded, the compositions preferably contain from about 0.1% to about10%, by weight compositions, of Matrixyl® and/or Biopeptide CL®peptide-containing compositions.

Anti-Oxidants/Radical Scavengers

The compositions of the present invention may include a safe andeffective amount of an anti-oxidant/radical scavenger. Theanti-oxidant/radical scavenger is especially useful for providingprotection against UV radiation which can cause increased scaling ortexture changes in the stratum corneum and against other environmentalagents which can cause skin damage.

A safe and effective amount of an anti-oxidant/radical scavenger may beadded to the compositions of the subject invention, preferably fromabout 0.1% to about 10%, more preferably from about 1% to about 5%, ofthe composition.

Anti-oxidants/radical scavengers such as ascorbic acid (vitamin C) andits salts, ascorbyl esters of fatty acids, ascorbic acid derivatives(e.g., magnesium ascorbyl phosphate, sodium ascorbyl phosphate, ascorbylsorbate), tocopherol (vitamin E), tocopherol sorbate, tocopherolacetate, other esters of tocopherol, butylated hydroxy benzoic acids andtheir salts, 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid(commercially available under the tradename Trolox.sup.R), gallic acidand its alkyl esters, especially propyl gallate, uric acid and its saltsand alkyl esters, sorbic acid and its salts, lipoic acid, amines (e.g.,N,N-diethylhydroxylamine, amino-guanidine), sulfhydryl compounds (e.g.,glutathione), dihydroxy fumaric acid and its salts, lycine pidolate,arginine pilolate, nordihydroguaiaretic acid, bioflavonoids, curcumin,lysine, methionine, proline, superoxide dismutase, silymarin, teaextracts, grape skin/seed extracts, melanin, and rosemary extracts maybe used. Preferred anti-oxidants/radical scavengers are selected fromtocopherol sorbate and other esters of tocopherol, more preferablytocopherol sorbate. For example, the use of tocopherol sorbate intopical compositions and applicable to the present invention isdescribed in U.S. Pat. No. 4,847,071, issued on Jul. 11, 1989 to DonaldL. Bissett, Rodney D. Bush and Ranjit Chatterjee.

Chelators

The compositions of the present invention may also contain a safe andeffective amount of a chelator or chelating agent. As used herein,“chelator” or “chelating agent” means an active agent capable ofremoving a metal ion from a system by forming a complex so that themetal ion cannot readily participate in or catalyze chemical reactions.The inclusion of a chelating agent is especially useful for providingprotection against UV radiation which can contribute to excessivescaling or skin texture changes and against other environmental agentswhich can cause skin damage.

A safe and effective amount of a chelating agent may be added to thecompositions of the subject invention, preferably from about 0.1% toabout 10%, more preferably from about 1% to about 5%, of thecomposition. Exemplary chelators that are useful herein are disclosed inU.S. Pat. No. 5,487,884, issued Jan. 30, 1996 to Bissett et al.;International Publication No. 91/16035, Bush et al., published Oct. 31,1995; and International Publication No. 91/16034, Bush et al., publishedOct. 31, 1995. Preferred chelators useful in compositions of the subjectinvention are furildioxime, furilmonoxime, and derivatives thereof.

Flavonoids

The compositions of the present invention may optionally contain aflavonoid compound. Flavonoids are broadly disclosed in U.S. Pat. Nos.5,686,082 and 5,686,367, both of which are herein incorporated byreference. Flavonoids suitable for use in the present invention areflavanones selected from unsubstituted flavanones, mono-substitutedflavanones, and mixtures thereof, chalcones selected from unsubstitutedchalcones, mono-substituted chalcones, di-substituted chalcones,tri-substituted chalcones, and mixtures thereof, flavones selected fromunsubstituted flavones, mono-substituted flavones, di-substitutedflavones, and mixtures thereof, one or more isoflavones; coumarinsselected from unsubstituted coumarins, mono-substituted coumarins,di-substituted coumarins, and mixtures thereof; chromones selected fromunsubstituted chromones, mono-substituted chromones, di-substitutedchromones, and mixtures thereof, one or more dicoumarols; one or morechromanones; one or more chromanols; isomers (e.g., cis/trans isomers)thereof, and mixtures thereof. By the term “substituted” as used hereinmeans flavonoids wherein one or more hydrogen atom of the flavonoid hasbeen independently replaced with hydroxyl, C₁-C₈ alkyl, C₁-C₄ alkoxyl,0-glycoside, and the like or a mixture of these substituents.

Examples of suitable flavonoids include, but are not limited to,unsubstituted flavanone, mono-hydroxy flavanones (e.g., 2′-hydroxyflavanone, 6-hydroxy flavanone, 7-hydroxy flavanone, etc.), mono-alkoxyflavanones (e.g., 5-methoxy flavanone, 6-methoxy flavanone, 7-methoxyflavanone, 4′-methoxy flavanone, etc.), unsubstituted chalcone(especially unsubstituted trans-chalcone), mono-hydroxy chalcones (e.g.,2′-hydroxy chalcone, 4′-hydroxy chalcone, etc.), di-hydroxy chalcones(e.g., 2′,4-dihydroxy chalcone, 2′,4′-dihydroxy chalcone, 2,2′-dihydroxychalcone, 2′,3-dihydroxy chalcone, 2′,5′-dihydroxy chalcone, etc.), andtri-hydroxy chalcones (e.g., 2′,3′,4′-trihydroxy chalcone,4,2′,4′-trihydroxy chalcone, 2,2′,4′-trihydroxy chalcone, etc.),unsubstituted flavone, 7,2′-dihydroxy flavone, 3′,4′-dihydroxynaphthoflavone, 4′-hydroxy flavone, 5,6-benzoflavone, and7,8-benzoflavone, unsubstituted isoflavone, daidzein (7,4′-dihydroxyisoflavone), 5,7-dihydroxy-4′-methoxy isoflavone, soy isoflavones (amixture extracted from soy), unsubstituted coumarin, 4-hydroxy coumarin,7-hydroxy coumarin, 6-hydroxy-4-methyl coumarin, unsubstituted chromone,3-formyl chromone, 3-formyl-6-isopropyl chromone, unsubstituteddicoumarol, unsubstituted chromanone, unsubstituted chromanol, andmixtures thereof.

Preferred for use herein are unsubstituted flavanone, methoxyflavanones, unsubstituted chalcone, 2′,4-dihydroxy chalcone, andmixtures thereof. More preferred are unsubstituted flavanone,unsubstituted chalcone (especially the trans isomer), and mixturesthereof.

They can be synthetic materials or obtained as extracts from naturalsources (e.g., plants). The naturally sourced material can also furtherbe derivatized (e.g., an ester or ether derivative prepared followingextraction from a natural source). Flavonoid compounds useful herein arecommercially available from a number of sources, e.g., Indofine ChemicalCompany, Inc. (Somerville, N.J.), Steraloids, Inc. (Wilton, N.H.), andAldrich Chemical Company, Inc. (Milwaukee, Wis.).

Mixtures of the above flavonoid compounds may also be used.

The herein described flavonoid compounds are preferably present in theinstant invention at concentrations of from about 0.01% to about 20%,more preferably from about 0.1% to about 10%, and still more preferablyfrom about 0.5% to about 5%.

Anti-Inflammatory Agents

A safe and effective amount of an anti-inflammatory agent may be addedto the compositions of the present invention, preferably from about 0.1%to about 10%, more preferably from about 0.5% to about 5%, of thecomposition. The anti-inflammatory agent enhances the skin appearancebenefits of the present invention, e.g., such agents contribute to amore uniform and acceptable skin tone or color. The exact amount ofanti-inflammatory agent to be used in the composition will depend on theparticular anti-inflammatory agent utilized since such agents varywidely in potency.

Steroidal anti-inflammatory agents, including but not limited to,corticosteroids such as hydrocortisone, hydroxyltriamcinolone,alpha-methyl dexamethasone, dexamethasone-phosphate, beclomethasonedipropionates, clobetasol valerate, desonide, desoxymethasone,desoxycorticosterone acetate, dexamethasone, dichlorisone, diflorasonediacetate, diflucortolone valerate, fluadrenolone, flucloroloneacetonide, fludrocortisone, flumethasone pivalate, fluosinoloneacetonide, fluocinonide, flucortine butylesters, fluocortolone,fluprednidene (fluprednylidene) acetate, flurandrenolone, halcinonide,hydrocortisone acetate, hydrocortisone butyrate, methylprednisolone,triamcinolone acetonide, cortisone, cortodoxone, flucetonide,fludrocortisone, difluorosone diacetate, fluradrenolone,fludrocortisone, difluorosone diacetate, fluradrenolone acetonide,medrysone, amcinafel, amcinafide, betamethasone and the balance of itsesters, chloroprednisone, chlorprednisone acetate, clocortelone,clescinolone, dichlorisone, diflurprednate, flucloronide, flunisolide,fluoromethalone, fluperolone, fluprednisolone, hydrocortisone valerate,hydrocortisone cyclopentylpropionate, hydrocortamate, meprednisone,paramethasone, prednisolone, prednisone, beclomethasone dipropionate,triamcinolone, and mixtures thereof may be used. The preferred steroidalanti-inflammatory for use is hydrocortisone.

A second class of anti-inflammatory agents which is useful in thecompositions of the present invention includes the nonsteroidalanti-inflammatory agents. The variety of compounds encompassed by thisgroup are well-known to those skilled in the art. For detaileddisclosure of the chemical structure, synthesis, side effects, etc. ofnon-steroidal anti-inflammatory agents, one may refer to standard texts,including Anti-inflammatory and Anti-Rheumatic Drugs, K. D. Rainsford,Vol. I-III, CRC Press, Boca Raton, (1985), and Anti-inflammatory Agents,Chemistry and Pharmacology, 1, R. A. Scherrer, et al., Academic Press,New York (1974).

Specific non-steroidal anti-inflammatory agents useful in thecomposition of the invention include, but are not limited to: 1) theoxicams, such as piroxicam, isoxicam, tenoxicam, sudoxicam, andCP-14,304; 2) the salicylates, such as aspirin, disalcid, benorylate,trilisate, safapryn, solprin, diflunisal, and fendosal; 3) the aceticacid derivatives, such as diclofenac, fenclofenac, indomethacin,sulindac, tolmetin, isoxepac, furofenac, tiopinac, zidometacin,acematacin, fentiazac, zomepirac, clindanac, oxepinac, felbinac, andketorolac; 4) the fenamates, such as mefenamic, meclofenamic,flufenamic, niflumic, and tolfenamic acids; 5) the propionic acidderivatives, such as ibuprofen, naproxen, benoxaprofen, flurbiprofen,ketoprofen, fenoprofen, fenbufen, indopropfen, pirprofen, carprofen,oxaprozin, pranoprofen, miroprofen, tioxaprofen, suprofen, alminoprofen,and tiaprofenic; and 6) the pyrazoles, such as phenylbutazone,oxyphenbutazone, feprazone, azapropazone, and trimethazone.

Mixtures of these non-steroidal anti-inflammatory agents may also beemployed, as well as the dermatologically acceptable salts and esters ofthese agents. For example, etofenamate, a flufenamic acid derivative, isparticularly useful for topical application. Of the nonsteroidalanti-inflammatory agents, ibuprofen, naproxen, flufenamic acid,etofenamate, aspirin, mefenamic acid, meclofenamic acid, piroxicam andfelbinac are preferred; ibuprofen, naproxen, ketoprofen, etofenamate,aspirin and flufenamic acid are more preferred.

Finally, so-called “natural” anti-inflammatory agents are useful in thecomposition of the present invention. Such agents may suitably beobtained as an extract by suitable physical and/or chemical isolationfrom natural sources (e.g., plants, fungi, by-products ofmicroorganisms) or can be synthetically prepared. For example,candelilla wax, bisabolol (e.g., alpha bisabolol), aloe vera, plantsterols (e.g., phytosterol), Manjistha (extracted from plants in thegenus Rubia, particularly Rubia Cordifolia), and Guggal (extracted fromplants in the genus Commiphora, particularly Commiphora Mukul), kolaextract, chamomile, red clover extract, and sea whip extract, may beused.

Additional anti-inflammatory agents useful herein include compounds ofthe Licorice (the plant genus/species Glycyrrhiza glabra) family,including glycyrrhetic acid, glycyrrhizic acid, and derivatives thereof(e.g., salts and esters). Suitable salts of the foregoing compoundsinclude metal and ammonium salts. Suitable esters include C₂-C₂₄saturated or unsaturated esters of the acids, preferably C₁₀-C₂₄, morepreferably C₁₆-C₂₄. Specific examples of the foregoing include oilsoluble licorice extract, the glycyrrhizic and glycyrrhetic acidsthemselves, monoammonium glycyrrhizinate, monopotassium glycyrrhizinate,dipotassium glycyrrhizinate, 1-beta-glycyrrhetic acid, stearylglycyrrhetinate, and 3-stearyloxy-glycyrrhetinic acid, and disodium3-succinyloxy-beta-glycyrrhetinate. Stearyl glycyrrhetinate ispreferred.

Anti-Cellulite Agents

The compositions of the present invention may also contain a safe andeffective amount of an anti-cellulite agent. Suitable agents mayinclude, but are not limited to, xanthine compounds (e.g., caffeine,theophylline, theobromine, and aminophylline).

Topical Anesthetics

The compositions of the present invention may also contain a safe andeffective amount of a topical anesthetic. Examples of topical anestheticdrugs include benzocaine, lidocaine, bupivacaine, chlorprocaine,dibucaine, etidocaine, mepivacaine, tetracaine, dyclonine, hexylcaine,procaine, cocaine, ketamine, pramoxine, phenol, and pharmaceuticallyacceptable salts thereof.

Tanning Actives

The compositions of the present invention may contain a tanning active.When present, it is preferable that the compositions contain from about0.1% to about 20%, more preferably from about 2% to about 7%, and stillmore preferably from about 3% to about 6%, by weight of the composition,of dihydroxyacetone as an artificial tanning active.

Dihydroxyacetone, which is also known as DHA or1,3-dihydroxy-2-propanone, is a white to off-white, crystalline powder.This material can be represented by the chemical formula C₃H₆O₃. Thecompound can exist as a mixture of monomers and dimers, with the dimerspredominating in the solid crystalline state. Upon heating or melting,the dimers break down to yield the monomers. This conversion of thedimeric form to the monomeric form also occurs in aqueous solution.Dihydroxyacetone is also known to be more stable at acidic pH values.See The Merck Index, Tenth Edition, entry 3167, p. 463 (1983), and“Dihydroxyacetone for Cosmetics”, E. Merck Technical Bulletin, 03-304110, 319 897, 180 588.

Skin Lightening Agents

The compositions of the present invention may contain a skin lighteningagent. When used, the compositions preferably contain from about 0.1% toabout 10%, more preferably from about 0.2% to about 5%, also preferablyfrom about 0.5% to about 2%, by weight of the composition, of a skinlightening agent. Suitable skin lightening agents include those known inthe art, including kojic acid, arbutin, ascorbic acid and derivativesthereof (e.g., magnesium ascorbyl phosphate, sodium ascorbyl phosphateor ascorbyl glucoside), and extracts (e.g., mulberry extract, placentalextract). Skin lightening agents suitable for use herein also includethose described in the PCT publication No. 95/34280, in the name ofHillebrand, corresponding to PCT Application No. U.S. 95/07432, filedJun. 12, 1995; and co-pending U.S. application Ser. No. 08/390,152 filedin the names of Kvalnes, Mitchell A. DeLong, Barton J. Bradbury, CurtisB. Motley, and John D. Carter, corresponding to PCT Publication No.95/23780, published Sep. 8, 1995.

Skin Soothing and Skin Healing Actives

The compositions of the present invention may comprise a skin soothingor skin healing active. Skin soothing or skin healing actives suitablefor use herein include panthenoic acid derivatives (including panthenol,dexpanthenol, ethyl panthenol), aloe vera, allantoin, bisabolol, anddipotassium glycyrrhizinate. A safe and effective amount of a skinsoothing or skin healing active may be added to the present composition,preferably, from about 0.1% to about 30%, more preferably from about0.5% to about 20%, still more preferably from about 0.5% to about 10%,by weight of the composition formed.

Antimicrobial and Antifungal Actives

The compositions of the present invention may contain an antimicrobialor antifungal active. Such actives are capable of destroying microbes,preventing the development of microbes or preventing the pathogenicaction of microbes. A safe and effective amount of an antimicrobial orantifungal active may be added to the present compositions, preferably,from about 0.001% to about 10%, more preferably from about 0.01% toabout 5%, and still more preferably from about 0.05% to about 2%.

Examples of antimicrobial and antifungal actives include B-lactam drugs,quinolone drugs, ciprofloxacin, norfloxacin, tetracycline, erythromycin,amikacin, 2,4,4′-trichloro-2′-hydroxy diphenyl ether,3,4,4′-trichlorobanilide, phenoxyethanol, phenoxy propanol,phenoxyisopropanol, doxycycline, capreomycin, chlorhexidine,chlortetracycline, oxytetracycline, clindamycin, ethambutol, hexamidineisethionate, metronidazole, pentamidine, gentamicin, kanamycin,lineomycin, methacycline, methenamine, minocycline, neomycin,netilmicin, paromomycin, streptomycin, tobramycin, miconazole,tetracycline hydrochloride, erythromycin, zinc erythromycin,erythromycin estolate, erythromycin stearate, amikacin sulfate,doxycycline hydrochloride, capreomycin sulfate, chlorhexidine gluconate,chlorhexidine hydrochloride, chlortetracycline hydrochloride,oxytetracycline hydrochloride, clindamycin hydrochloride, ethambutolhydrochloride, metronidazole hydrochloride, pentamidine hydrochloride,gentamicin sulfate, kanamycin sulfate, lineomycin hydrochloride,methacycline hydrochloride, methenamine hippurate, methenaminemandelate, minocycline hydrochloride, neomycin sulfate, netilmicinsulfate, paromomycin sulfate, streptomycin sulfate, tobramycin sulfate,miconazole hydrochloride, ketaconazole, amanfadine hydrochloride,amanfadine sulfate, octopirox, parachlorometa xylenol, nystatin,tolnaftate, zinc pyrithione and clotrimazole.

Preferred examples of actives useful herein include those selected fromsalicylic acid, benzoyl peroxide, 3-hydroxy benzoic acid, glycolic acid,lactic acid, 4-hydroxy benzoic acid, acetyl salicylic acid,2-hydroxybutanoic acid, 2-hydroxypentanoic acid, 2-hydroxyhexanoic acid,cis-retinoic acid, trans-retinoic acid, retinol, phytic acid,N-acetyl-L-cysteine, lipoic acid, azelaic acid, arachidonic acid,benzoylperoxide, tetracycline, ibuprofen, naproxen, hydrocortisone,acetominophen, resorcinol, phenoxyethanol, phenoxypropanol,phenoxyisopropanol, 2,4,4′-trichloro-2′-hydroxy diphenyl ether,3,4,4′-trichlorocarbanilide, octopirox, lidocaine hydrochloride,clotrimazole, miconazole, ketoconazole, neocycin sulfate, and mixturesthereof.

Sunscreen Actives

Exposure to ultraviolet light can result in excessive scaling andtexture changes of the stratum corneum. Therefore, the composition ofthe present invention may optionally contain a sunscreen active. As usedherein, “sunscreen active” includes both sunscreen agents and physicalsunblocks. Suitable sunscreen actives may be organic or inorganic.

Inorganic sunscreen actives useful herein include the following metallicoxides; titanium dioxide having an average primary particle size of fromabout 15 nm to about 100 nm, zinc oxide having an average primaryparticle size of from about 15 nm to about 150 nm, zirconium oxidehaving an average primary particle size of from about 15 nm to about 150nm, iron oxide having an average primary particle size of from about 15nm to about 500 nm, and mixtures thereof. When used herein, theinorganic sunscreen actives are present in the amount of from about 0.1%to about 20%, preferably from about 0.5% to about 10%, more preferablyfrom about 1% to about 5%, by weight of the composition.

A wide variety of conventional organic sunscreen actives are suitablefor use herein. Sagarin, et al., at Chapter VIII, pages 189 et seq., ofCosmetics Science and Technology (1972), discloses numerous suitableactives. Specific suitable sunscreen actives include, for example:p-aminobenzoic acid, its salts and its derivatives (ethyl, isobutyl,glyceryl esters; p-dimethylaminobenzoic acid); anthranilates (i.e.,o-amino-benzoates; methyl, menthyl, phenyl, benzyl, phenylethyl,linalyl, terpinyl, and cyclohexenyl esters); salicylates (amyl, phenyl,octyl, benzyl, menthyl, glyceryl, and di-pro-pyleneglycol esters);cinnamic acid derivatives (menthyl and benzyl esters, a-phenylcinnamonitrile; butyl cinnamoyl pyruvate); dihydroxycinnamic acidderivatives (umbelliferone, methylumbelliferone,methylaceto-umbelliferone); trihydroxy-cinnamic acid derivatives(esculetin, methylesculetin, daphnetin, and the glucosides, esculin anddaphnin); hydrocarbons (diphenylbutadiene, stilbene); dibenzalacetoneand benzalacetophenone; naphtholsulfonates (sodium salts of2-naphthol-3,6-disulfonic and of 2-naphthol-6,8-disulfonic acids);di-hydroxynaphthoic acid and its salts; o- andp-hydroxybiphenyldisulfonates; coumarin derivatives (7-hydroxy,7-methyl, 3-phenyl); diazoles (2-acetyl-3-bromoindazole, phenylbenzoxazole, methyl naphthoxazole, various aryl benzothiazoles); quininesalts (bisulfate, sulfate, chloride, oleate, and tannate); quinolinederivatives (8-hydroxyquinoline salts, 2-phenylquinoline); hydroxy- ormethoxy-substituted benzophenones; uric and violuric acids; tannic acidand its derivatives (e.g., hexaethylether); (butyl carbotol) (6-propylpiperonyl)ether; hydroquinone; benzophenones (oxybenzene, sulisobenzone,dioxybenzone, benzoresorcinol, 2,2′,4,4′-tetrahydroxybenzophenone,2,2′-dihydroxy-4,4′-dimethoxybenzophenone, octabenzone;4-isopropyldibenzoylmethane; butylmethoxydibenzoylmethane; etocrylene;octocrylene; [3-(4′-methylbenzylidene bornan-2-one), terephthalylidenedicamphor sulfonic acid and 4-isopropyl-di-benzoylmethane.

Of these, 2-ethylhexyl-p-methoxycinnamate (commercially available asPARSOL MCX), 4,4′-t-butyl methoxydibenzoyl-methane (commerciallyavailable as PARSOL 1789), 2-hydroxy-4-methoxybenzophenone,octyldimethyl-p-aminobenzoic acid, digalloyltrioleate,2,2-dihydroxy-4-methoxybenzophenone,ethyl-4-(bis(hydroxy-propyl))aminobenzoate,2-ethylhexyl-2-cyano-3,3-diphenylacrylate, 2-ethylhexyl-salicylate,glyceryl-p-aminobenzoate, 3,3,5-tri-methylcyclohexylsalicylate,methylanthranilate, p-dimethyl-aminobenzoic acid or aminobenzoate,2-ethylhexyl-p-dimethyl-amino-benzoate, 2-phenylbenzimidazole-5-sulfonicacid, 2-(p-dimethylaminophenyl)-5-sulfonicbenzoxazoic acid, octocryleneand mixtures of these compounds, are preferred.

More preferred organic sunscreen actives useful in the compositions are2-ethylhexyl-p-methoxycinnamate, butylmethoxydibenzoyl-methane,2-hydroxy-4-methoxybenzo-phenone, 2-phenylbenzimidazole-5-sulfonic acid,octyldimethyl-p-aminobenzoic acid, octocrylene and mixtures thereof.

Also particularly useful in the compositions of the invention aresunscreen actives such as those disclosed in U.S. Pat. No. 4,937,370issued to Sabatelli on Jun. 26, 1990, and U.S. Pat. No. 4,999,186 issuedto Sabatelli & Spirnak on Mar. 12, 1991. The sunscreening agentsdisclosed therein have, in a single molecule, two distinct chromophoremoieties which exhibit different ultra-violet radiation absorptionspectra. One of the chromophore moieties absorbs predominantly in theUVB radiation range and the other absorbs strongly in the UVA radiationrange.

Preferred members of this class of sunscreening agents are4-N,N-(2-ethylhexyl)methyl-aminobenzoic acid ester of2,4-dihydroxybenzophenone; N,N-di-(2-ethylhexyl)-4-aminobenzoic acidester with 4-hydroxydibenzoylmethane;4-N,N-(2-ethylhexyl)methyl-aminobenzoic acid ester with4-hydroxydibenzoylmethane; 4-N,N-(2-ethylhexyl)methyl-aminobenzoic acidester of 2-hydroxy-4-(2-hydroxyethoxy)benzophenone;4-N,N-(2-ethylhexyl)-methylaminobenzoic acid ester of4-(2-hydroxyethoxy)dibenzoylmethane;N,N-di-(2-ethylhexyl)-4-aminobenzoic acid ester of2-hydroxy-4-(2-hydroxyethoxy)benzophenone; andN,N-di-(2-ethylhexyl)-4-aminobenzoic acid ester of4-(2-hydroxyethoxy)dibenzoylmethane and mixtures thereof.

Especially preferred sunscreen actives include4,4′-t-butylmethoxydibenzoylmethane, 2-ethylhexyl-p-methoxycinnamate,phenyl benzimidazole sulfonic acid, and octocrylene.

A safe and effective amount of the organic sunscreen active is used,typically from about 1% to about 20%, more typically from about 2% toabout 10% by weight of the composition. Exact amounts will varydepending upon the sunscreen or sunscreens chosen and the desired SunProtection Factor (SPF).

Particulate Material

The compositions of the present invention may contain a particulatematerial, preferably a metallic oxide. These particulates can be coatedor uncoated, charged or uncharged. Charged particulate materials aredisclosed in U.S. Pat. No. 5,997,887, to Ha, et al., incorporated hereinby reference. Particulate materials useful herein include; bismuthoxychloride, iron oxide, mica, mica treated with barium sulfate andTiO2, silica, nylon, polyethylene, talc, styrene, polypropylene,ethylene/acrylic acid copolymer, sericite, titanium dioxide, bismuthoxychloride, iron oxide, aluminum oxide, silicone resin, barium sulfate,calcium carbonate, cellulose acetate, polymethyl methacrylate, andmixtures thereof.

Inorganic particulate materials, e.g., TiO2, ZnO, or ZrO2 arecommercially available from a number of sources. One example of asuitable particulate material contains the material available from U.S.Cosmetics (TRONOX TiO2 series, SAT-T CR837, a rutile TiO2). Preferably,particulate materials are present in the composition in levels of fromabout 0.01% to about 2%, more preferably from about 0.05% to about 1.5%,still more preferably from about 0.1% to about 1%, by weight of thecomposition.

Conditioning Agents

The composition of the present invention may contain a conditioningagent selected from humectants, moisturizers, or skin conditioners. Avariety of these materials can be employed and each can be present at alevel of from about 0.01% to about 20%, more preferably from about 0.1%to about 10%, and still more preferably from about 0.5% to about 7% byweight of the composition. These materials include, but are not limitedto, guanidine; urea; glycolic acid and glycolate salts (e.g. ammoniumand quaternary alkyl ammonium); salicylic acid; lactic acid and lactatesalts (e.g., ammonium and quaternary alkyl ammonium); aloe vera in anyof its variety of forms (e.g., aloe vera gel); polyhydroxy alcohols suchas sorbitol, mannitol, xylitol, erythritol, glycerol, hexanetriol,butanetriol, propylene glycol, butylene glycol, hexylene glycol and thelike; polyethylene glycols; sugars (e.g., melibiose) and starches; sugarand starch derivatives (e.g., alkoxylated glucose, fucose, glucosamine);hyaluronic acid; lactamide monoethanolamine; acetamide monoethanolamine;panthenol; allantoin; and mixtures thereof. Also useful herein are thepropoxylated glycerols described in U.S. Pat. No. 4,976,953, to Orr etal, issued Dec. 11, 1990.

Also useful are various C₁-C₃₀ monoesters and polyesters of sugars andrelated materials. These esters are derived from a sugar or polyolmoiety and one or more carboxylic acid moieties. Such ester materialsare further described in, U.S. Pat. No. 2,831,854, U.S. Pat. No.4,005,196, to Jandacek, issued Jan. 25, 1977; U.S. Pat. No. 4,005,195,to Jandacek, issued Jan. 25, 1977, U.S. Pat. No. 5,306,516, to Letton etal, issued Apr. 26, 1994; U.S. Pat. No. 5,306,515, to Letton et al,issued Apr. 26, 1994; U.S. Pat. No. 5,305,514, to Letton et al, issuedApr. 26, 1994; U.S. Pat. No. 4,797,300, to Jandacek et al, issued Jan.10, 1989; U.S. Pat. No. 3,963,699, to Rizzi et al, issued Jun. 15, 1976;U.S. Pat. No. 4,518,772, to Volpenhein, issued May 21, 1985; and U.S.Pat. No. 4,517,360, to Volpenhein, issued May 21, 1985.

Preferably, the conditioning agent is selected from urea, guanidine,sucrose polyester, panthenol, dexpanthenol, allantoin, and combinationsthereof.

Structuring Agents

The compositions hereof, and especially the emulsions hereof, maycontain a structuring agent. Structuring agents are particularlypreferred in the oil-in-water emulsions of the present invention.Without being limited by theory, it is believed that the structuringagent assists in providing rheological characteristics to thecomposition which contribute to the stability of the composition. Forexample, the structuring agent tends to assist in the formation of theliquid crystalline gel network structures. The structuring agent mayalso function as an emulsifier or surfactant. Preferred compositions ofthis invention contain from about 0.1% to about 20%, more preferablyfrom about 0.1% to about 10%, still more preferably from about 0.5% toabout 9%, of one or more structuring agents.

Preferred structuring agents are those having an HLB of from about 1 toabout 8 and having a melting point of at least about 45° C. Suitablestructuring agents are those selected from saturated C₁₄ to C₃₀ fattyalcohols, saturated C₁₆-C₃₀ fatty alcohols containing from about 1 toabout 5 moles of ethylene oxide, saturated C₁₆-C₃₀ diols, saturatedC₁₆-C₃₀ monoglycerol ethers, saturated C₁₆-C₃₀ hydroxy fatty acids,C₁₄-C₃₀ hydroxylated and nonhydroxylated saturated fatty acids, C₁₄-C₃₀saturated ethoxylated fatty acids, amines and alcohols containing fromabout 1 to about 5 moles of ethylene oxide diols, C₁₄-C₃₀ saturatedglyceryl mono esters with a monoglyceride content of at least 40%,C₁₄-C₃₀ saturated polyglycerol esters having from about 1 to about 3alkyl group and from about 2 to about 3 saturated glycerol units,C₁₄-C₃₀ glyceryl mono ethers, C₁₄-C₃₀ sorbitan mono/diesters, C₁₄-C₃₀saturated ethoxylated sorbitan mono/diesters with about 1 to about 5moles of ethylene oxide, C₁₄-C₃₀ saturated methyl glucoside esters,C₁₄-C₃₀ saturated sucrose mono/diesters, C₁₄-C₃₀ saturated ethoxylatedmethyl glucoside esters with about 1 to about 5 moles of ethylene oxide,C₁₄-C₃₀ saturated polyglucosides having an average of between 1 to 2glucose units and mixtures thereof, having a melting point of at leastabout 45° C.

The preferred structuring agents of the present invention are selectedfrom stearic acid, palmitic acid, stearyl alcohol, cetyl alcohol,behenyl alcohol, stearic acid, palmitic acid, the polyethylene glycolether of stearyl alcohol having an average of about 1 to about 5ethylene oxide units, the polyethylene glycol ether of cetyl alcoholhaving an average of about 1 to about 5 ethylene oxide units, andmixtures thereof. More preferred structuring agents of the presentinvention are selected from stearyl alcohol, cetyl alcohol, behenylalcohol, the polyethylene glycol ether of stearyl alcohol having anaverage of about 2 ethylene oxide units (steareth-2), the polyethyleneglycol ether of cetyl alcohol having an average of about 2 ethyleneoxide units, and mixtures thereof. Even more preferred structuringagents are selected from stearic acid, palmitic acid, stearyl alcohol,cetyl alcohol, behenyl alcohol, steareth-2, and mixtures thereof.

Thickening Agent (Including Thickeners and Gelling Agents)

The compositions of the present invention can contain one or morethickening agents, preferably from about 0.1% to about 5%, morepreferably from about 0.1% to about 4%, and still more preferably fromabout 0.25% to about 3%, by weight of the composition.

Nonlimiting classes of thickening agents include those selected from thefollowing:

a) Carboxylic Acid Polymers

These polymers are crosslinked compounds containing one or more monomersderived from acrylic acid, substituted acrylic acids, and salts andesters of these acrylic acids and the substituted acrylic acids, whereinthe crosslinking agent contains two or more carbon-carbon double bondsand is derived from a polyhydric alcohol. Polymers useful in the presentinvention are more fully described in U.S. Pat. No. 5,087,445, to Haffeyet al, issued Feb. 11, 1992; U.S. Pat. No. 4,509,949, to Huang et al,issued Apr. 5, 1985; U.S. Pat. No. 2,798,053, to Brown, issued Jul. 2,1957; and in CTFA International Cosmetic Ingredient Dictionary, FourthEdition, 1991, pp. 12 and 80.

Examples of commercially available carboxylic acid polymers usefulherein include the carbomers, which are homopolymers of acrylic acidcrosslinked with allyl ethers of sucrose or pentaerytritol. Thecarbomers are available as the Carbopol® 900 series from B.F. Goodrich(e.g., Carbopol® 954). In addition, other suitable carboxylic acidpolymeric agents include copolymers of C₁₀₋₃₀ alkyl acrylates with oneor more monomers of acrylic acid, methacrylic acid, or one of theirshort chain (i.e., C₁₋₄ alcohol) esters, wherein the crosslinking agentis an allyl ether of sucrose or pentaerytritol. These copolymers areknown as acrylates/C₁₀₋₃₀ alkyl acrylate crosspolymers and arecommercially available as Carbopol® 1342, Carbopol® 1382, Pemulen TR-1,and Pemulen TR-2, from B.F. Goodrich. In other words, examples ofcarboxylic acid polymer thickeners useful herein are those selected fromcarbomers, acrylates/C₁₀-C₃₀ alkyl acrylate crosspolymers, and mixturesthereof.

b) Crosslinked Polyacrylate Polymers

The compositions of the present invention can optionally containcrosslinked polyacrylate polymers useful as thickeners or gelling agentsincluding both cationic and nonionic polymers, with the cationics beinggenerally preferred. Examples of useful crosslinked nonionicpolyacrylate polymers and crosslinked cationic polyacrylate polymers arethose described in U.S. Pat. No. 5,100,660, to Hawe et al, issued Mar.31, 1992; U.S. Pat. No. 4,849,484, to Heard, issued Jul. 18, 1989; U.S.Pat. No. 4,835,206, to Farrar et al, issued May 30, 1989; U.S. Pat. No.4,628,078 to Glover et al issued Dec. 9, 1986; U.S. Pat. No. 4,599,379to Flesher et al issued Jul. 8, 1986; and EP 228,868, to Farrar et al,published Jul. 15, 1987.

c) Polyacrylamide Polymers

The compositions of the present invention can optionally containpolyacrylamide polymers, especially nonionic polyacrylamide polymersincluding substituted branched or unbranched polymers. More preferredamong these polyacrylamide polymers is the nonionic polymer given theCTFA designation polyacrylamide and isoparaffin and laureth-7, availableunder the Tradename Sepigel 305 from Seppic Corporation (Fairfield,N.J.).

Other polyacrylamide polymers useful herein include multi-blockcopolymers of acrylamides and substituted acrylamides with acrylic acidsand substituted acrylic acids. Commercially available examples of thesemulti-block copolymers include Hypan SR150H, SS500V, SS500W, SSSA100H,from Lipo Chemicals, Inc., (Patterson, N.J.).

d) Polysaccharides

A wide variety of polysaccharides are useful herein. “Polysaccharides”refer to gelling agents which contain a backbone of repeating sugar(i.e., carbohydrate) units. Nonlimiting examples of polysaccharidegelling agents include those selected from cellulose, carboxymethylhydroxyethylcellulose, cellulose acetate propionate carboxylate,hydroxyethylcellulose, hydroxyethyl ethylcellulose,hydroxypropylcellulose, hydroxypropyl methylcellulose, methylhydroxyethylcellulose, microcrystalline cellulose, sodium cellulosesulfate, and mixtures thereof. Also useful herein are the alkylsubstituted celluloses. In these polymers, the hydroxy groups of thecellulose polymer is hydroxyalkylated (preferably hydroxyethylated orhydroxypropylated) to form a hydroxyalkylated cellulose which is thenfurther modified with a C₁₀-C₃₀ straight chain or branched chain alkylgroup through an ether linkage. Typically these polymers are ethers ofC₁₀-C₃₀ straight or branched chain alcohols with hydroxyalkylcelluloses.Examples of alkyl groups useful herein include those selected fromstearyl, isostearyl, lauryl, myristyl, cetyl, isocetyl, cocoyl (i.e.alkyl groups derived from the alcohols of coconut oil), palmityl, oleyl,linoleyl, linolenyl, ricinoleyl, behenyl, and mixtures thereof.Preferred among the alkyl hydroxyalkyl cellulose ethers is the materialgiven the CTFA designation cetyl hydroxyethylcellulose, which is theether of cetyl alcohol and hydroxyethylcellulose. This material is soldunder the tradename Natrosol® CS Plus from Aqualon Corporation(Wilmington, Del.).

Other useful polysaccharides include scleroglucans which are a linearchain of (1-3) linked glucose units with a (1-6) linked glucose everythree units, a commercially available example of which is Clearogel™CS11 from Michel Mercier Products Inc. (Mountainside, N.J.).

e) Gums

Other thickening and gelling agents useful herein include materialswhich are primarily derived from natural sources. Nonlimiting examplesof these gelling agent gums include acacia, agar, algin, alginic acid,ammonium alginate, amylopectin, calcium alginate, calcium carrageenan,carnitine, carrageenan, dextrin, gelatin, gellan gum, guar gum, guarhydroxypropyltrimonium chloride, hectorite, hyaluroinic acid, hydratedsilica, hydroxypropyl chitosan, hydroxypropyl guar, karaya gum, kelp,locust bean gum, natto gum, potassium alginate, potassium carrageenan,propylene glycol alginate, sclerotium gum, sodium carboxymethyl dextran,sodium carrageenan, tragacanth gum, xanthan gum, and mixtures thereof.

Preferred compositions of the present invention include a thickeningagent selected from carboxylic acid polymers, crosslinked polyacrylatepolymers, polyacrylamide polymers, and mixtures thereof, more preferablyselected from carboxylic acid polymers, polyacrylamide polymers, andmixtures thereof.

Preferably, the extract containing phenylethanoid glycosides isextracted from the dry stems of Cistanche tubulosa (Schenk.) Wight.

According to the invention, a process for preparing the extractcontaining phenylethanoid glycosides from Cistanche tubulosa (Schenk.)is provided. The process comprises the steps of:

a) Grinding the dry stem of cistanche tubulosa (Schenk.) Wight to form acrude powder;

b) Soaking the crude powder with water or ethanol, and refluxing themixture at a temperature of 50-100° C. and collecting the supernant;

c) Concentrating the supernatant obtained in step b) at a decreasedpressure and centrifuging the concentrate;

d) Adding the centrifugate obtained in step c) onto a column which ispacked with a macro-porous resin SEPABEADA;

e) Eluting the column with 10-60% of ethanol or deionized water;

f) Collecting the eluant and subjecting it to be dried under vacuum,crushed and sieved to obtain an extract containing phenylethanoidglycosides.

The extraction of Cistanche tubulosa (Schenk.) Wight includes twoprocedures:

The primary extraction procedure includes soaking with water or ethanol,boiling, filtering, cooling, precipitating and centrifuging. Thecentrifugate obtained is the crude preparation. Preferably, therefluxing in step b) is repeated 2-4 times and the temperature ispreferably from 70 to 90° C.

The refining procedure includes adding the centrifugate onto the columnpacked with a macro-porous resin, then adsorbing and eluting with agradient concentration of ethanol or deionized water. The eluantobtained is a refining preparation, which is further dried under vacuum,crashed and sieved to obtain an extract containing phenylethanoidglycosides.

In combination with the above-mentioned carriers or adjuvants, theextract from Cistanche tubulosa (Schenk.) Wight can be formulated toobtain various cosmetics, such as facial mask, lotion, facial cream andointment, and dermic preparations. These cosmetics and dermicpreparations possess effects of overall skin improvement in softness,moisture, smoothness, whitening and spot reduction.

Preparation of the Cosmetic Composition

The Cosmetic skin care compositions of the present invention aregenerally prepared by using conventional methods such as those known inthe art. Such methods typically involve mixing of the ingredients in oneor more steps to a relatively uniform state, with or without heating,cooling, application of vacuum, and the like.

Methods for Regulating Skin Condition

The current Cosmetic compositions are useful in regulating mammalianskin. The preferred method for application is via leave-on products suchas skin gels or skin creams. A less preferred method for application isvia rinse-off or wipe-off products such as cleansers, both foamingcleansers and cold cream cleansers.

Regulating mammalian skin condition involves topically applying to thekeratinous tissue a safe and effective amount of a composition of thepresent invention. The amount of the composition which is applied, thefrequency of application and the period of use will vary widelydepending upon the level of the skin care active substance and/or othercomponents of a given composition and the level of regulation desired,e.g., in light of the level of keratinous tissue damage present orexpected to occur.

In a preferred embodiment, the composition is chronically applied to theskin. By “chronic topical application” is meant continued topicalapplication of the composition over an extended period during thesubject's lifetime, preferably for a period of at least about one week,more preferably for a period of at least about one month, even morepreferably for at least about three months, even more preferably for atleast about six months, and more preferably still for at least about oneyear. While benefits are obtainable after various maximum periods of use(e.g., five, ten or twenty years), it is preferred that chronicapplication continue throughout the subject's lifetime. Typicallyapplications would be on the order of about once per day over suchextended periods, however application rates can vary from about once perweek up to about three times per day or more.

A wide range of quantities of the compositions of the present inventioncan be employed to provide a skin appearance and/or feel benefit.Quantities of the present compositions which are typically applied perapplication are, in mg composition/cm² skin, from about 0.1 mg/cm² toabout 10 mg/cm². A particularly useful application amount is about 1mg/cm² to about 2 mg/cm².

Regulating mammalian skin tissue condition is preferably practiced byapplying a composition in the form of a skin lotion, cream, gel, foam,ointment, paste, emulsion, spray, conditioner, tonic, cosmetic,lipstick, foundation, nail polish, after-shave, or the like which ispreferably intended to be left on the skin or other keratin structurefor some esthetic, prophylactic, therapeutic or other benefit (i.e., a“leave-on” composition). After applying the composition to the skin, itis preferably left on the skin for a period of at least about 15minutes, more preferably at least about 30 minutes, even more preferablyat least about 1 hour, still more preferably for at least several hours,e.g., up to about 12 hours. Any part of the external portion of theface, hair, and/or nails can be treated, e.g., face, lips, under-eyearea, eyelids, scalp, neck, torso, arms, hands, legs, feet, fingernails,toenails, scalp hair, eyelashes, eyebrows, etc. The composition can beapplied with the fingers or with an implement or device (e.g., pad,cotton ball, applicator pen, spray applicator, and the like).

Another approach to ensure a continuous exposure of the skin to at leasta minimum level of the plant extract and skin care active is to applythe compound by use of a patch applied, e.g., to the face. Such anapproach is particularly useful for problem skin areas needing moreintensive treatment (e.g., facial crows feet area, frown lines, undereye area, and the like). The patch can be occlusive, semi-occlusive ornon-occlusive and can be adhesive or non-adhesive. The composition canbe contained within the patch or be applied to the skin prior toapplication of the patch. The patch can also include additional activessuch as chemical initiators for exothermic reactions such as thosedescribed in U.S. Pat. Nos. 5,821,250, 5,981,547, and 5,972,957 to Wu,et al. The patch is preferably left on the skin for a period of at leastabout 5 minutes, more preferably at least about 10 minutes, morepreferably still at least about 15 minutes, even more preferably atleast about 20 minutes, still more preferably at night as a form ofnight therapy.

The benefits of the invention: Various skin care cosmetic compositionswere prepared by blending an extract from Cistanche deserticola,especially Cistanche tubulosa (schenk.) Wight, as an essential activesubstance with other adjuvants and/or additives. No toxicity or sideeffect has been shown in acute and chronic toxicological experiments inanimals, and no allergic response has been shown in clinical tests,either. In addition, the tyrosinase-inhibiting activity and the actionon the capillary permeability of the cosmetic composition of theinvention are several times of commercial freckle-removing cream. Thefinished products of the cosmetic composition of the invention showevident effects upon applying onto the human skin for several weeks. Ithas been shown in a series of tests that the extract from Cistanchetubulosa (schenk.) Wight possesses skin-whitening, freckle-removing andbrightening effects, and has no toxicity or side effect to the skin.Even no stimulation has been shown in the skin. Thus, the cosmeticcomposition of the invention has a practical applicability in theindustry.

PREFERRED EMBODIMENTS Preparation of the Plant Extract from cistanchetubulosa (Schenk) Wight Example 1

10 kg of cistanche tubulosa (schenk.) wight was crashed into a crudepowder, then soaked in 50% ethanol in an amount which was 8 times of thecrude powder. The mixture was heated at a temperature above 50° C. fortwo hours. The mixture was filtered through a 100 mesh screen. Theresidue was then extracted twice with 50% ethanol in an amount which was6 times of the residue, and the extraction was filtered through a 100mesh screen. The combined filtrate was concentrated under vacuum (thedegree of vacuum≧0.08 Mpa) such that the concentrate has a specificgravity of 1.10. The concentrate was cooled and precipitated for 12hours. The supernatant was centrifuged and the centrifugate was addedonto a column packed with SEPABEADS resin. The column was eluted withtwo volume of 20% ethanol and the eluant was collected. The column waseluted twice, with two volume of 30% ethanol each time. The eluants ofthe total three times were combined and concentrated at a reducedpressure to obtain an extrative containing phenylethanoid glycosides andhaving a weight of 710 g.

The contents of acteoside and echinacoside were measured via a highperformance liquid chromatography (HPLC), with the stationary phasebeing silicone of C18 alkyl silane; the mobile phase beingacetonitrile-methanol-1% acetic acid (10:15:75) aqueous solution in aflow rate of 1 ml/min, and the detecting wavelength being 334 nm.

Acteoside and echinacoside were weighed precisely and dissolved in themobile phase to prepare reference solutions, with each 1 ml solutioncontaining 0.1 mg acteoside and 0.14 mg echinacoside, respectively.

The preparation of the test solution: 0.5 g of extract was added in a 50ml brown measuring flask, into which 25 ml of the mobile phase was addedprecisely. The flask was weighted precisely and kept standing for 0.5hr. After being ultrasonicated (230 W, 35 kHz, 40 min), the flask wascooled and weighted again. The lost weight was made up with the mobilephase. Then, the flask was shaken, centrifuged and kept standing. Thesupernatant was removed to another brown bottle.

Measurement: The reference solution and the test solution were taken outin an amount of 2-10 μl and 10-20 μl, respectively, and injected intothe liquid chromatography column. The peak areas of the acteoside andthe echinacoside were measured. The contents were calculated from thepeak areas. The content of the acteoside is 9.6% while the content ofthe echinacoside is 5.2% by weight of the preparation.

Example 2

10 kg of cistanche tubulosa (schenk.) wight was crashed into a crudepowder, then soaked in 50% ethanol in an amount which was 8 times of thecrude powder. The mixture was heated for two hours, then filteredthrough a 100 mesh screen. The residue was heated twice each with 50%ethanol in an amount which was 6 times of the residue, and the mixturewas filtered through a 100 mesh screen. The combined filtrate wasconcentrated under vacuum (the degree of vacuum≧0.08 Mpa) such that theconcentrate has a specific gravity of 1.10. The concentrate was cooledand precipitated for 12 hours. The supernatant was centrifuged and thecentrifugate was added onto a column packed with SEPABEADS resin. Thecolumn was eluted with two volume of 20% ethanol and the eluant wascollected. Then the column was eluted with two volume of 30% ethanol.The later eluting step were repeated twice. The eluants of the totalfour times were combined and concentrated at a reduced pressure toobtain an extrative containing phenylethanoid glycosides and having aweight of 490 g. The content of the acteoside is 21.3% while the contentof the echinacoside is 9.2% by weight of the preparation according tothe above measurement.

Example 3

10 kg of cistanche tubulosa (schenk.) wight was crashed into a crudepowder, then soaked in 50% ethanol in an amount which was 8 times of thecrude powder. The mixture was heated for two hours, then filteredthrough a 100 mesh screen. The residue was heated three times each with50% ethanol in an amount which was 6 times of the residue, and themixture was filtered through a 100 mesh screen. The combined filtratewas concentrated under vacuum (the degree of vacuum≧0.08 Mpa) such thatthe concentrate has a specific gravity of 1.10. The concentrate wascooled and precipitated for 12 hours. The supernatant was centrifugedand the centrifugate was added onto a column packed with SEPABEADSresin. The column was eluted with two volume of 30% ethanol and theeluant was collected. Then the column was eluted with two volume of 40%ethanol. The later eluting step were repeated three times. The eluantsof the total five times were combined and concentrated at a reducedpressure to obtain an extrative containing phenylethanoid glycosides andhaving a weight of 160 g. The content of the acteoside is 65.3% whilethe content of the echinacoside is 22.2% by weight of the preparationaccording to the above measurement.

When the cistanche tubulosa (schenk.) wight was extracted with water,the contents (wt %) of acteoside and echinacoside were higher than thosewhen the extraction is undertaken with an alcohol.

Pharmacological effects: Skin melanocytes located in the basal layer ofepidermis are the basis of the synthesis of melanine in the humanpegmentary system. The melanine is synthesized from tyrosineenzymatically by tyrosinase in melanocytes and deposits in melanosomesto form uniform melanin granules gradually. The activity of thetyrosinase influences directly the formation of the melanine. It hasbeen shown in experiments that the inhibition of phenylethanoidglycosides on the activity of tyrosinase is significantly stronger thanprunol, the well-known brightening agent used in the cosmetics (p>0.05).

It has been observed, in tissue culture experiments, that the cytolergyof M cells form a collagen fiber network, and acteoside, aphenylethanoid glycoside, possesses an obvious promotion on the growthof M cells, so as to accelerate the formation of the collagen fibernetwork. The difference in comparison with the control group isstatistically significant (P>0.05).

Experimental Example 1. Acute Toxicity (LD₅₀, p.o.) Experiment

Purpose: Observation on the adverse reaction and death conditionresulted from the administration of test samples in a single dose, p.o.,and determination of the half-lethal dose

Basis of detection: GB15193.3-1994

1.1. Materials and Method:

1.1.1 Sample: cistanche tubulosa (schenk.) wight

1.1.2 Characters of the sample: Deep brown powder

1.1.3 Preparation of the sample: Sample 10000 mg was weighted andgrinded. Distilled water was added into the grinded sample to 20 ml, andthe mixture was blended sufficiently to form a uniform suspension whichwas used as a test sample.

1.1.4 Animal subject: 20 Kunming species of mice, 18-22 g, half male andhalf female, provided by the experimental animal department, FudanUniversity. Certificate No.: 02-22-1. Temperature of bleeding room:18-22° C. Relative humidity of bleeding room: 40-70%. Certificate No. ofanimal cabinet: 02-28. The feedstuffs were provided by SuhangExperimental Animal Tech-developing Co. and the Certificate No. offeedstuffs is Su E Shi Shen (2002) 006.

1.1.5 Instrumentation: Electronic scale AC-3A 90401582, Equi-armbalanceBP3100S-91006909.

1.1.6 Test methods:

1.1.6.1 After fasting (no refraining from water), the mice were selectedbased on the body weight, 10 male and 10 female, and housed into twocabinets, respectively. The difference of body weight is below 3 g amongthe same sex mice.

1.1.6.2 Each animal was weighted and administrated with a single dose ofthe test sample in a volume of 0.4 ml/20 g via an intragastric catheter.

1.1.6.3 The general conditions, the change of weight, the toxic symptomsand the death conditions were observed for one week.

1.1.6.4 The animals were weighted again at the end of the experiment.The dead animals and the animals sacrificed at the end of the experimentwere necrotomied and the pathologic changes were detectedmacroscopically.

1.1.6.5 The whole course and observed conditions were recorded indetail.

1.2 Results:

TABLE 1 Results of acute toxicity test in mice (p.o.) Dose Animal DeathFatality Sexuality (mg/kg) (N) (n) (%) Female 10000 10 0 0 Male 10000 100 0

1.2.1 Cardinal symptoms: The activities of animals in each group arenormal and the capills are in good color and glossiness during the wholeexperimental stage. No toxic symptom or death was observed.

1.2.2 Half-lethal dose: LD₅₀>10000 mg/kg, both in female and male mice.

1.3 Conclusion: The test sample is substantially an asepsis substanceaccording to the grading on the half-lethal dose.

2. Micronucleus Test

Purpose: Detection of marrow chromosomal aberration induced in mice invivo

Basis of detection: GB-15193.5-1994

Sample: Cistanche tubulosa (schenk.) wight

2.1 Characters of the sample: Extract from Cistanche tubulosa (schenk.)wight, Deep brown powder

2.2 Preparation of sample: Sample 5000, 2500 or 1250 mg was weighted andgrinded. Distilled water was added into the grinded sample to 20 ml, andthe mixture was blended sufficiently to form a test sample.

2.3 Animal subject: Kunming species of mice, 25-30 g, provided byexperimental animal department, Fudan University. Certificate No.:02-22-1.

2.4 Test condition: Temperature: 18-22° C. Relative humidity: 40-70%.

2.5 Test methods:

2.5.1 Animals were divided randomly into 5 groups, 10 in each group withhalf of female and half of male. In the five groups, there were threegroups for three doses of test sample, one negative control group,distilled water, and one positive control group, cyclophosphaide.

2.5.2. Animals were weighted and administrated with the test sample orcontrol substances in different concentrations in a volume of 0.4 ml/20g via an intragastric catheter, and the administration is repeated oncemore in 30 hr.

2.5.3 Animals were sacrificed via cervical bertebra dislocation 6 hr.after the second intragastric administration. The marrow was taken outof the femoral bone and blended uniformly with calf serum. Then, thefilm preparation, the fixation, and Giemasa stain were undertakenconventionally.

2.5.4 Test under microscope: Numbers of micronuclei were counted in overone thousand of philo-pleochromatic akaryocytes. Incidence rates ofmicronuclei were calculated and analyzed statistically.

2.6 Results:

TABLE 2 Incidence rates of micronucle in philo-pleochromatic akaryocytesin animal marrow Rate of Dose Micronucleated Micronuclei StatisticTest(mg/kg) Sex No. Cells tested Cells No. (‰) (p value) 5000 M 5 5000 40.8 >0.05 5000 F 5 5000 3 0.6 >0.05 2500 M 5 5000 5 1.0 >0.05 2500 F 55000 3 0.6 >0.05 1250 M 5 5000 4 0.8 >0.05 1250 F 5 5000 4 0.8 >0.05Dis. water M 5 5000 4 0.8 (20000 mg/kg) F 5 5000 4 0.8 Cyclophosphaide M5 5000 132 26.4 <0.01 (40 mg/kg) F 5 5000 129 25.8 <0.01 *Compared withthe negative contral group (chi square test).

2.7 Conclusion: Negative results of the Extract from Cistanche tubulosa(schenk.) wight. have been shown statistically in the micronucleus testof eosinophil cells in marrow.

3. Teratospermia Test

Purpose: Detection of the influence of the test sample on thegonepoiesis and the development of spermatozoa, and detection of thegenetoxic on the cell granule in vivo

Basis of detection: GB-15193.7-1994

3.1 Characters of the sample: Extract from Cistanche tubulosa (schenk.)wight, Deep brown powder

3.2 Preparation of sample: Sample 5000, 2500 or 1250 mg was weighted andgrinded. Distilled water was added into the grinded sample to 20 ml, andthe mixture was blended sufficiently to obtain a test sample.

3.3 Animal subject: Male mice, depuratory grade, 25-30 g, provided byexperimental animal department, Fudan University. Certificate No.:02-22-1.

3.4 Test condition: Temperature: 18-22° C. Relative humidity: 40-70%.

3.5 Test methods:

3.5.1 Animals were divided randomly into 5 groups, 5 in each group. Itthe five groups, there were three groups for three doses of test sample,one negative control group, distilled water, and one positive controlgroup, cyclophosphaide.

3.5.2. Animals were weighted and administrated with the test sample orcontrol substances in different concentrations in a volume of 0.4 ml/20g via an intragastric catheter, once a day for 5 days.

3.5.3 Animals were sacrificed via cervical bertebra dislocation 35 daysafter the first intragastric administration. The ambo-epididymises weretaken off and put into normal saline. After being cut into pieces, theywere filtered through a four layer gauze. The filter was smeared, fixedand dyed with 2% eosin.

3.5.4 Test under microscope: Amounts of teratosperm in 1000 spernatozoaswere counted per mouse and rates of teratospermia were calculatedstatistically.

3.6 Results:

TABLE 3 Rates of teratospermia in mice Type of aberration (n) DoseSpernatozoa No Banana Fat Fold Double Double (mg/kg) No. tested crooksharp head amorphism tail head tail 5000 5 5000 2 1 3 48 0 0 0 2500 55000 3 2 4 51 0 0 0 1250 5 5000 3 1 4 48 0 0 0 Dis. water 5 5000 3 2 549 0 0 0 (20000 mg/kg) Cyclophosphaide 5 5000 28 32 45 175 2 4 6 (40mg/kg) Dose An-spermatozoa (mg/kg) (n) Aberration rate (‰) Statistictest (p) 5000 54 10.8 >0.05 2500 60 12.0 >0.05 1250 56 11.2 >0.05 Dis.water 59 11.8 (20000 mg/kg) Cyclophosphaide 292 58.4 <0.01 (40 mg/kg)*Compared with the negative contral (chi square test).

4. AmesTest

3.7 Results: Negative results have been shown statistically in theteratospermia test of the Extract from Cistanche tublosa (schenk.)wight.

Purpose: Detection of Mutagenicity of the test substance to predict itsgenetic injury and potential carcinogenesis

Basis of detection: GB15193.4-1994

4.1 Characters of the sample: Extract from Cistanche tubulosa (schenk.)wight, Deep brown powder

4.2 Solvent: Sterile distilled water

4.3 Sample processing and preparation: 4000 mg sample was weighted anddistilled water was added into the sample to 20 ml, then the mixture wasblended to obtain a 200 mg/ml of solution. 3, 1, 0.3 and 0.1 mL of thissolution were taken, respectively and each was added with steriledistilled water to 10 ml to obtain a solution in the concentration of 60mg/ml, 20 mg/ml, 6 mg/ml and 2 mg/ml, respectively.

4.4 Test doses: 0.2 mg/dish, 0.6 mg/dish, 2 mg/dish, 6 mg/dish and 20mg/dish

4.5 Environic condition of the experiment: Temperature: 18-22° C.,Relative humidity: 50-70%

4.6 Instrumentation: Incubator PYX-DHS 193

-   -   Thermostatic water bath DK-600    -   Electronic balance AE163 1010002

4.7 Test strains: TA97, TA98, TA100 and TA102, with qualitied biologicalcharacters, provided by Biochemistry Dept., California University, CA,America. The concentration of the bacteria liquid used for the test was1-2×10⁹/ml.

4.8 Induction and preparation of rat liver S₉:

A healthy adult SD rat with a body weight of about 150 g was selected.Polychlorinatedbiphenyl was dissolved into corn oil to obtain a solutionof 200 mg/ml. The above solution was injected (i.p.), in a sterileoperation, into the rat in a dose of 500 mg/kg. 5 days later, the animalwas sacrificed via decapitation. The liver was taken out and weighted,then rinsed several times with 0.15 mol/L of fresh cooled solution ofpotassium chloride. The amount of the potassium chloride solution was 3ml/g liver (wet weight). The beaker in which the rinsed liver put wasremoved into an ice bath, and the liver was cut into pieces with a pairof sterilized scissors. The liver was homogenized with a tissuehomogenizer (2000 r/min, 1 min), then the liver homogenate thus preparedwas centrifuged in a high speed centrifuge (0-4° C., 9000 g, 10 min).The supernatant, S₉ part, was packed into several sterile tubes andconserved in liquid nitrogen. The above steps need to be operated in asterile and cooled local circumstance. The S₉ thus prepared was furtheridentified for its content of protein and bioactivity of indirectmutagen. Result of identification: the sterility test was qualified, thecontent of protein was 33 mg/ml, and the S₉ bioactivity conformed withthe criterion (0.5 ml S₉ mixture/dish, in which 50 μl of S₉ wascontained).

4.9 Solvent control: Sterilee distilled water

4.10 Positive control:

−S₉: TA97 Atebrin (500 μg/dish), TA98 p-nitroquinoline (200 μg/dish),TA100 (1 μl/dish), TA102 (1 μl/dish), and MMS (1 μl/dish).

+S₉: TA97, TA98, TA100 2-aminofluorene (10 μg/dish), TA102 1,8-dioxyanthraquinone (50 μl/dish).

The above positive control substances were dissolved in dimethylsulfoxide except Atebrin which was dissolved in sterile distilled water.

4.11 Method of test:

Standard plate incorporation on the basis of GB15193-94: Into the topagar (45° C., 2 ml), the bacterial liquid 0.1 ml, Sample 0.1 ml and S9mixture 0.5 ml for activation were added sequently. The mixture wasblended sufficiently and poured rapidly onto the underlayer medium.After the incubation of 48 hr. at 37° C., results were observed. eachtest was repeated once more.

4.12 Results of the first test:

TABLE 4 Re-established colonies (colonies/sish) X ± SD Dose TA97 TA98TA100 TA102 (mg/dish) −S₉ +S₉ −S₉ +S₉ −S₉ +S₉ −S₉ +S₉  0.2 147 ± 7 154 ±3 35 ± 4 37 ± 5 162 ± 8 164 ± 6  246 ± 10 249 ± 7  0.6 150 ± 5 148 ± 636 ± 2 35 ± 3 164 ± 6 161 ± 4 244 ± 8  251 ± 11  2.0 153 ± 4 150 ± 4 34± 5 36 ± 4 158 ± 7 163 ± 7 251 ± 8 246 ± 7  6.0 146 ± 6 147 ± 8 37 ± 335 ± 3 165 ± 6 159 ± 7 243 ± 7 245 ± 9 20.0 141 ± 5 148 ± 8 36 ± 4 38 ±2 157 ± 5 163 ± 8 236 ± 7  240 ± 10 Blank 148 ± 6 154 ± 4 36 ± 3 37 ± 5163 ± 7 158 ± 8  247 ± 10 250 ± 6 Solvent 151 ± 7 148 ± 5 35 ± 4 34 ± 4161 ± 6 165 ± 4 245 ± 8 246 ± 7 Atebrin  1626 ± 109 (500 μg/dish)p-nitroquinoline 1108 ± 73  (200 μg/dish) Methyl 1463 ± 84  2784 ± 289mesylate (1 μl/dish) 2-aminofluorene 1727 ± 64 2196 ± 143  1618 ± 118(10 μg/dish) 1,8-dioxy  885 ± 56 anthraquinone (50 μl/dish)

4.13 Results of the second test:

TABLE 5 Re-established colonies (colonies/sish) X ± SD Dose TA97 TA98TA100 TA102 (mg/dish) −S₉ +S₉ −S₉ +S₉ −S₉ +S₉ −S₉ +S₉  0.2 139 ± 5 142 ±7 40 ± 5 41 ± 3 146 ± 7 149 ± 8 267 ± 8 260 ± 9  0.6 136 ± 6 145 ± 6 39± 3 40 ± 3 150 ± 6 151 ± 4 264 ± 7  266 ± 11  2.0 142 ± 4 145 ± 7 38 ± 338 ± 5 148 ± 4 149 ± 7  267 ± 10 246 ± 6  6.0 137 ± 8 138 ± 8 41 ± 2 41± 6 145 ± 5 143 ± 6 258 ± 4 261 ± 9 20.0 135 ± 6 140 ± 6 37 ± 4 38 ± 3142 ± 5 150 ± 5 259 ± 8 256 ± 6 Blank 136 ± 6 144 ± 5 39 ± 5 40 ± 4 147± 6 149 ± 3 264 ± 9 270 ± 7 Solvent 143 ± 6  41 ± 3 43 ± 3 144 ± 7  146± 5 267 ± 6 268 ± 4 143 ± 6 Atebrin 1478 ± 47 (500 μg/dish)p-nitroquinoline 868 ± 60 (200 μg/dish) Methyl 1641 ± 80  2520 ± 112mesylate (1 μl/dish) 2-aminofluorene  1772 ± 137 2774 ± 164  1548 ± 124(10 μg/dish) 1,8-dioxy  975 ± 86 anthraquinone (50 μl/dish)

4.14 Results: Negative results have been shown in the Extract fromCistanche tubulosa (schenk.) wight. in the above test condition.

5. 30 day Feeding trial

Purpose: Further detection on the toxic action of the sample andestimation of the maximum non-toxic dose

Basis of detection: GB15193.7-1994

5.1 Name of the sample: Cistanche tubulosa (schenk.) wight

5.2 Characters of the sample: Deep brown powder

5.3 Design of doses: The recommended dose for human body is 1080 mg/60kg/d. There were three dose groups, 180 mg/kg (high), 900 mg/kg (middle)and 1800 mg/kg (low), corresponding to 10 times, 50 times and 100 timesof the recommended dose for human body, respectively, and a blankcontrol group.

5.4 Sample processing:

According to the dose design of the test and the feed intaken by theanimal (about 10 g/100 g BW/d), 18, 90 and 180 g of sample wereincorporated into 10 kg of feed, respectively, mixed thoroughly, andgranula feedstuffs containing high, middle or low amount of sample wereproduced. Three groups of animals were feeded with these three dosefeedstuffs. The blank control group of animals were feeded with thesimilar granula feedstuffs except having no sample.

5.5 Experimental animals and the circumstance:

5.5.1 80 SD rats, 60-80 g, half male and half female, provided byexperimental animal department, Fudan University. Certificate No.:02-22-2.

5.5.2 Temperature of bleeding: 18-22° C. Relative humidity: 40-70%.Certificate No. of animal cabinet: 02-28. The feedstuffs were providedby Suhang Experimental Animal Tech-developing Co. and the CertificateNo. of feedstuffs is Su E Shi Shen (2002) 006.

5.6 Instrumentation:

Automatic biochemical equipment AL800 (Shimadzu, Japan) 15R325

Blood analyser CD3700 (Cell-DYN America Corporation) 15R325

Desk centrifuge Mikro22 (HETTTCH Company) 1105

Electronic balance Bp3100s (Sand Sartorius Corporation) 91006823

Electronic balance JA1003 (Shanghai Balance Factory) 300

Electronic scale ACS-3 (Weighing Apparatus Factory, Shanghai Wo) 1050480

Electronic scale ACS-3 (Weighing Apparatus Factory, Shanghai Wo)90401570

5.7 Method of test:

5.7.1 Animals were divided randomly into four groups, 20 for each group,half male and half female, three groups for three doses, respectively,and one group for the blank control. The animals were kept alone, withfood and drink, ad lib., for 30 days.

5.7.2 The body weights were weighed and the intake of the foodstuffswere recorded at the beginning of the test and once a week in the wholeperiod of test.

5.7.3 30 Days later, rats were weighed and blood samples were collectedfor the hematologic and biochemical analysis. Then rats were sacrificed,dissected and observed macroscopically to detect any obviouspathological changes. The liver, kidney, spleen and sex organs, etc.,were weighed for the calculation of the ratio of organ weight to bodyweight, and the liver, kidney, spleen, gastro-intestine and sex organs,etc. were histopathologically inspected

Results:

5.8.1 Growth status and food availability:

TABLE 6 Growth of body weight of experimental animals (g) ( X ± SD) SexGroup 0 week 1 week 2 weeks 3 weeks 4 weeks ♂ Control 71 ± 6  122 ± 10177 ± 12 228 ± 15 279 ± 19 Low-dose 70 ± 6  120 ± 10 175 ± 12 224 ± 17282 ± 23 Intermindiat-dose 71 ± 7  122 ± 10 178 ± 13 226 ± 18 284 ± 24High-dosage 71 ± 6 119 ± 9 178 ± 14 227 ± 19 276 ± 24 ♀ Control 71 ± 7113 ± 9 150 ± 11 171 ± 13 292 ± 13 Low-dose 71 ± 7 114 ± 9 148 ± 10 171± 12 188 ± 17 Intermindiat-dose 70 ± 7 114 ± 9 147 ± 11 169 ± 12 186 ±16 High-dosage 72 ± 6 114 ± 8 148 ± 11 170 ± 13 184 ± 15

TABLE 7 Food availiability of experimental animals ( X ± SD) Food SexGroup Beginning (g) End (g) Growth (g) Ingestion (g) availiability (%) ♂Control 71 ± 6 279 ± 19 208 ± 15 643 ± 39 32.3 ± 1.2 Low dose 70 ± 6 282± 23 212 ± 17 632 ± 49 33.6 ± 1.8 Intermindiat 71 ± 7 284 ± 24 213 ± 17673 ± 43 31.6 ± 1.0 dose High dosage 71 ± 6 276 ± 24 205 ± 19 632 ± 6832.5 ± 1.5 ♀ Control 71 ± 7 192 ± 13 121 ± 9  524 ± 49 23.3 ± 1.7 Lowdose 71 ± 7 188 ± 17 117 ± 12 512 ± 54 22.9 ± 1.7 Intermindiat 70 ± 7186 ± 16 116 ± 11 530 ± 57 21.8 ± 1.4 dose High dosage 72 ± 6 184 ± 15112 ± 10 498 ± 42 22.7 ± 1.9

It is shown in the above two tables that the growth of rats in eachsample group is essentially fine.

TABLE 8 Gross anotomy and macroscopic observation in kidney Group *Highdose Blank Animal 20 20 Cortical Renal capsule changing(cases) 0 0 partRenal tubular Necrosis(cases) 0 0 epithelial cell Edema(cases) 0 0denaturali- Vacuolated changes 0 0 zation (hydropic degeneration andfatty degeneration (cases) Cast nephropathy(cases) 4 5 Inflammatory cell0 0 infiltration(cases) Glomerular denaturalization(cases) 0 0Tubulointerstitial lesion(inflammatory 0 0 cell infiltration) (cases)Others(cases) 0 0 Renal Renal tubule Necrosis(cases) 0 0 medulladenaturali- Edema(cases) 0 0 zation Vacuolated changes 1 0 (hydropicdegeneration and fatty degeneration (cases) Cast nephropathy(cases) 0 0Inflammatory cell 0 0 infiltration(cases) Tubulointerstitial 0 0inflammatory cell infiltration (cases) Others(cases) 0 0 Renal pelvicpapillary changes(cases) 0 0 Transitional epithelium changes(cases) 0 0*It has been shown, in the gross anotomy and macroscopic observation,that the kidneys of rat in each sample group have no obviouspathological change, thus only rats in high dose group and contral groupwere histologically inspected.

TABLE 9 Gross anotomy and macroscopic observation in spleen Blank *Highdose Female Male Female Male Animal (N) 10 10 10 10 Red pulpextention(cases) 0 0 0 0 White pulp atrophy(cases) 0 0 0 0 Hematopoieticcell 0 0 0 0 hyperplasia(cases) Inflammatory cells 0 0 0 0infiltrate(cases) Hyperpigmentation(cases) 0 0 0 0 Others(cases) 0 0 0 0*It has been shown, in the gross anotomy and macroscopic observation,that the spleens of rat in each sample group have no obviouspathological change, thus only rats in high dose group and contral groupwere histologically inspected.

TABLE 10 Gross anotomy and macroscopic observation in gastro-intestineBlank *High dose Female Male Female Male Animal (N) 10 10 10 10 Mucosalbleeding (cases) 0 0 0 0 Edema(cases) 0 0 0 0 Inflammatory cells 0 0 1 0infiltrate(cases) Necrosis(cases) 0 0 0 0 Atrophy (cases) 0 0 0 0Proliferation (cases) 0 0 0 0 *It has been shown, in the gross anotomyand macroscopic observation, that the gastro-intestine of rat in eachsample group has no obvious pathological change, thus only rats in highdose group and contral group were histologically inspected.

TABLE 11 Gross anotomy and macroscopic observation in didymus Blank*High dose Animal (N) 10 10 Atrophy(cases) 0 0 Denaturalization (cases)0 0 Bleeding(cases) 0 0 Edema (cases) 0 0 Cysts(cases) 0 1 Others(cases)0 0 *It has been shown, in the gross anotomy and macroscopicobservation, that the didymus of rat in each sample group has no obviouspathological change, thus only rats in high dose group and contral groupwere histologically inspected.

TABLE 12 Gross anotomy and macroscopic observation in cvaries Blank*High dose Animal (N) 10 10 Atrophy(cases) 0 0 Denaturalization (cases)0 0 Bleeding(cases) 0 0 Edema (cases) 0 0 Cysts(cases) 2 1 Others(cases)0 0 *It has been shown, in the gross anotomy and macroscopicobservation, that Ovaries of rat in each sample group have no obviouspathological change, thus only rats in high dose group and contral groupwere histologically inspected.

The sample, the extract from Cistanche tubulosa (schenk.) wight, wastoxicological evaluated according to the stipulations in GB15193-1994,the results are as follows [(Inspec. No. 5970B, Tox. (2003) 0034),Preventive Medicine Research Center, Shanghai (2003)]:

1. Acute toxicity test: LD₅₀ (p.o.)>10000 mg/kg, both in female and malemice. The test sample is substantially an asepsis substance according tothe grading on the half-lethal dose.

2. Micronucleus Test: negative results.

3. Teratospermia Test: negative results.

4. AmesTest: negative results.

5. 30 Day Feeding Trial: Aminals in each group were well grown. There isno obvious difference between the sample group and the control group inthe results of hematologic analysis, biochemical analysis, ratio of mainorgan weight to body weight and histologic inspection.

6. Stimulation Test:

1) 6 Guinea pigs, 250±10 g, half male and half female, the skin wasshaved on two sides of vertebra 24 hr. prior to the administration ofsample, and the shaved area was 8×5 cm². The skin was inspected 24 hr.after shaving to ensure that no skin destruction exists.

Subject contrast was adopted on the left side and right side. The leftshaved side was applied with 1 g of the sample containing 2% of theextract from Cistanche tubulosa (schenk.) wight and other cosmeticadjuvants, and the right side was applied with 1 g of excipients as acontrol. The skin of each side was covered with a film, gauzes and fixedwith adhesive tapes. The guinea pigs were kept alone. 24 hr. later, thetest sample was washed-out with warm water. Then the areas of skin weremacroscopically observed and histopathologically inspected 1 hr., 24hr., 48 hr. and 72 hr., respectively, after the washing-out of the testsample. Also, the conditions of the position applied with the testsample were recorded and scored, such as erythema and/or oedema, if any.The scores and the recovery of above conditions were recorded every day.The average scores are listed in the following table for the evaluationof the stimulation of the test sample on the integrated skin (Table 13).

TABLE 13 Evaluation of stimulation on integrated skin Drug Control DrugControl No. Sex Weight (g) erythema oedema erythema oedema erythemaoedema erythema oedema 1 hr 24 hr 1 ♀ 245 1 0 0 0 0 0 0 0 2 ♀ 251 0 0 00 0 0 0 0 3 ♀ 259 0 0 0 0 0 0 0 0 4 ♂ 253 0 0 0 0 0 0 0 0 5 ♂ 249 0 0 00 0 0 0 0 6 ♂ 256 1 0 0 0 0 0 0 0 Average total scores 0.33 0 0 0Stimulation No stimulation 48 hr 72 hr 1 ♀ 245 0 0 0 0 0 0 0 0 2 ♀ 251 00 0 0 0 0 0 0 3 ♀ 259 0 0 0 0 0 0 0 0 4 ♂ 253 0 0 0 0 0 0 0 0 5 ♂ 249 00 0 0 0 0 0 0 6 ♂ 256 0 0 0 0 0 0 0 0 Average total scores 0 0 0 0Stimulation No stimulation

2) 6 Guinea pigs, 250±10 g, half male and half female, the skin wasshaved on two sides of vertebra 24 hr. prior to the administration ofsample, and the shaved area was 8×5 cm². The skin was sterilized andscarificated with a Scalpel to form a “#” scarification in a depth suchthat capillary hemorrhage can be seen. The damaged degree of skin isidentical essentially on both sides.

Subject contrast was adopted on the left side and right side. The leftshaved side was applied with 1 g of the sample containing 2% of theextract from Cistanche tubulosa (schenk.) wight and other cosmeticadjuvants, and the right side was applied with Ig of excipients as acontrol. The skin of each side was covered with a film, gauzes and fixedwith adhesive tapes. The guinea pigs were kept alone. 24 hr. later, thetest sample was washed-out with warm water. Then the skins weremacroscopically observed and histopathologically inspected 1 hr., 24hr., 48 hr. and 72 hr., respectively, after the washing-out of the testsample. Also, the conditions of the locations applied with the testsample were recorded and scored, such as erythema and/or oedema, if any.The scores and the recovery of above conditions were recorded every day.The average scores are listed in the following table 11 for theevaluation of the stimulation of the test sample on the damaged skin(Table 14).

TABLE 14 Evaluation of stimulation on damaged skin Drug Control DrugControl No. Sex Weight (g) erythema oedema erythema oedema erythemaoedema erythema oedema 1 hr 24 hr 1 ♀ 254 0 0 0 0 0 0 0 0 2 ♀ 258 1 0 00 0 0 0 0 3 ♀ 247 0 0 0 0 0 0 0 0 4 ♂ 252 0 0 0 0 0 0 0 0 5 ♂ 249 0 0 00 0 0 0 0 6 ♂ 255 0 0 0 0 0 0 0 0 Average total scores 0.33 0 0.33 0Stimulation No stimulation 48 hr 72 hr 1 ♀ 254 0 0 0 0 0 0 0 0 2 ♀ 258 00 0 0 0 0 0 0 3 ♀ 247 0 0 0 0 0 0 0 0 4 ♂ 252 0 0 0 0 0 0 0 0 5 ♂ 249 00 0 0 0 0 0 0 6 ♂ 255 0 0 0 0 0 0 0 0 Average total scores 0 0 0 0Stimulation No stimulation

7. Capillary Permeability Test

42 Mice were divided randomly into two groups, i.e., the experimentalgroup, the cosmetic composition of the invention containing 2% of theextract from Cistanche tubulosa (schenk.) wight and other cosmeticadjuvants; and the control group, the commercially available frecklecream containing arbutin as a main active. The abdomen of each mouse wasunhaired with a depilat. 36 hr. later, the cosmetic composition of theinvention or the commercially available freckle cream was applied on theabdomen of each mouse, 0.25 g/10 g, twice, with an interval of 20 min.Then, 0.02 ml of histamine solution was injected intracutaneously on theabdomen, at 1-2 position(s) per mouse. 0.2 ml of 1% Evans Blue solutionwas injected from the vena caudalis during 1-2 min. The mice weresacrificed via cervical vertebra luxatio 15 min.later. The abdomen skinwas splited and the blued area was measured with a caliper rule.

TABLE 15 Group Animal (N) Blued area (mm²) Cosmetic comosition of theinvention 21 87.3 ± 8.2 Commercially available freckle cream 21 56.2 ±4.9

8. Inhibition on tyrosinase

Materials: Tyrosinase available from Sigma; 3,4-dihydroxyphenylalanineContent>98%, BR (Shanghai Chemical Agent Factory); the cosmeticcomposition of the invention; and the commercially available frecklecream.

Experimental method: The cosmetic composition of the inventioncontaining 2% of the extract from Cistanche tubulosa (schenk.) wight andother cosmetic adjuvants and the commercially available freckle creamcontaining arbutin as a main active, both in the form of 50% methanolextract, were suspensed in a phosphate buffer (pH 6.8). This suspensionswere diluted with the buffer to desired concentrations for use.

Determination of the enzyme activity: According to the Method describedby Mason & Nagatsu, a mixture of 0.1 ml tyrosinase solution (1.375 μl),0.9 ml phosphate buffer and 1 ml buffer containing or not containing theextract from Cistanche tubulosa (schenk.) wight was incubated (25° C.,10 min), then 1 ml of 0.03% 3,4-Dihydroxyphenylalanine was added and themixture was incubated (25° C., 2 min). The incubation mixture wasdetermined in 475 nm to obtain the absorbance, then the inhibitionpercentage on the tyrosinase was calculated. 50% Inhibitionconcentration (ID₅₀) was used to evaluate the inhibition of the extractfrom Cistanche tubulosa (schenk.) wight on the tyrosinase (Table 16).

TABLE 16 Inhibition Concentration Dose Tyrosinase ratio Cosmeticcomposition 2.9 mg/ml 1 ml 1 ml 72.1% of the invention Commerciallyavailable 2.9 mg/ml 1 ml 1 ml 16.5% freckle cream

9. Application on Human Body

Subjects:

The cosmetic composition made from 2% of the extract from Cistanchetubulosa (schenk.) wight, as an active component, was applied to 60woman, 18-65 years old, to observe the freckle-removing and whiteningeffects. The applied sites were those with locus niger, colored spot,age pigmen, moth-patch, aestates, macula solaris, butterfly spot andcyasma, etc.

Application method: The cosmetic composition was applied uniformly onthe facial skin and left dorsum of hand without a time limit for theremaining, 2-4 times per day, for 2-4 weeks.

Judgment on Effects:

Obvious amelioration: Colored spots formed due to the accrescence ofskin pigment resulting from various factors almost disappear or may onlybe seen indistinctly; the skin of dorsum of the left hand was obviouspure white, exquisite, slick and flexible in comparison with the skin ofdorsum of the right hand.

Amelioration: Various colored spots were lightened, more than 30% ofcolored spots faded away, the skin of dorsum of the left hand was whiteand exquisite comparing with that of the right hand.

Inefficacy: No change in colored spots, the skin of dorsum of both handswere in an equal color.

Results: There were 16 obvious amelioration cases; 35 ameliorationcases; and 9 inefficacy cases. The total effective rate was 85%.

It is shown that effective rate of the cosmetic composition of theinvention is 85% and the effects are obvious within 2-4 weeks after theconventional application in a normal dose.

Preparation of the Cosmetic Composition of the Invention

Examples 1˜6

The following topical skin care compositions containing the plantextract from cistanche tubulosa (Schenk) wight. The process for thepreparation of these compositions are similar to those used for thepreparation of common cosmetic composition. The weight percents ofacteoside and echinacoside in the extract from cistanche tubulosa(Schenk) wight are 9.6% and 7.2%, 27.1% and 11.0%, 71.3% and 8.2%, 37.9%and 23.5%, 8.7% and 5.2%, and 75.1% and 12.2%, respectively, in Formula1-6.

TABLE 17 Formula 1 2 3 4 5 6 Extract from 2.0 1.0 0.5 1.0 2.0 2.0cistanche tublosa (Schenk) wight Deionized water 71.65 65.45 63.7 61.2561.0 58.34 Xanthan Gum 0.3 0 0 0 0 0 Glycerin 5.0 0 0 12.0 12.0 12.0Propylene Glycol 3.0 0 0 0 0 0 Disodium EDTA 0.2 0 0 0.2 0.2 0.2Carbopol #934 0.3 0.2 0.2 0 0 0 Isopropyl 2.4 0 2.0 0 0 0 PalmitateCetyl Alcohol 1.5 3.0 3.0 0.55 0.6 0.55 Ethylhexyl 4.0 0 0 0 0 0palmitate Methyl paraben 0.4 0.3 0.3 0.2 0.3 0.2 Tocopheryl acetate 0.10 0.1 0 0 0 Vitamin A 0.1 0 0.1 0 0 0 palmitate Petrolatum 2.0 6.0 5.0 00 0 Ethylene glycol 0.5 1.5 1.0 0 0 0 distearate Stearyl alcohol 2.71.25 2.0 0 0 0 ethoxylated (20-40) Triethanolamine 0.3 0.2 0.2 0 0 0Cyclomethicone 2.0 3.0 2.0 0 0 0 D5 Cyclomethicone 1.0 1.0 2.0 0 0 0Blend Germall 0.5 0 0 0 0 0 D&C Yellow 10 0.05 0.05 0.05 0 0 0Hydroxyethyl 0 0.3 0.3 0 0 0 cellulose Mineral oil 0 14.3 15.0 0 0 0Myristyl myristate 0 2.0 2.0 0 0 0 Butylated 0 0.05 0.06 0 0 0hydroxytoluene Imidazolidinyl 0 0.3 0.5 0 0 0 urea Almond oil 0 0.1 0 00 0 Potassium 0 0 0 2.42 2.5 2.4 hydroxide 45% Stearic acid 0 0 0 18.015.0 18.0 Silicone fluid 0 0 0 2.0 2.0 2.0 SF-96 Ethylhexyl 0 0 0 1.251.25 1.25 methoxycinnamate Benzyl alcohol 0 0 0 0.5 0.5 0.5 LicoriceExtract 0 0 0 0.1 0.1 0 PT-40 Butylene glycol 0 0 0 0.5 0.5 0.5 Colorand dye 0 0 0 0.065 0.05 0.06 Ascorbyl 0 0 0 0 2.0 0 glucoside Titaniumdioxide 1.0 Zinc oxide 1.0

It is shown, from the above experiment Example, that the cosmeticcomposition of the invention possesses freckle-removing, skin-whiteningand brightening effects, which are many times better than the congenerproducts available commercially. The cosmetic composition of theinvention has no toxicity or side effect to human body and skin.

1. A cosmetic composition comprising an effective amount of extract fromcistanche tubulosa (Schenk) Wight and other useful cosmetic adjuvant(s).2. The cosmetic composition according to claim 1, wherein said extractfrom Cistanche tubulosa (Schenk) Wight is extracted from the dry stemsof Cistanche tubulosa (Schenk.) Wight.
 3. The cosmetic compositionaccording to claim 1, wherein said extract of cistanche tubulosa(Schenk) Wight comprises at least 2% of acteoside by weight and at least4% of echinaoside by weight, based on the total weight of the extract.4. The cosmetic composition according to claim 3, wherein said extractof cistanche tubulosa (Schenk) Wight comprises 2-80% of acteoside byweight, and 4-80% of echinaoside by weight, based on the total weight ofthe extract.
 5. The cosmetic composition according to claim 1, whereinsaid cosmetic composition is in a form of facial mask, lotion, facialcream, ointment or cleaning solution.
 6. A process for preparing acosmetic composition with an effective amount of extract from cistanchetubulosa (Schenk) Wight and other useful cosmetic adjuvant(s) comprisingsteps of preparing said extract from cistanche tubulosa (Schenk) Wight:a. grinding the dry stem of cistanche tubulosa (Schenk.) Wight to form adry powder; b. Soaking said dry powder of cistanche tubulosa (Schenk)Wight with water or alcohol and refluxing the mixture at a temperatureof 50-100° C.; c. Concentrating the supernatant obtained in step b undera decreased pressure and centrifuging the concentrate thus obtained; d.adding the centrifugate obtained in step c onto a column which is packedwith a macro-porous resin such as SEPABEADA; e. eluting said column with10-60% of alcohol or deionized water; and f. Collecting the eluant andsubjecting it to be dried under vacuum, crushed and sieved to obtain anextract containing phenylethanoid glycosides.
 7. The process accordingto claim 6, wherein said refluxing in step b is repeated 2-4 times andthe temperature is from 70 to 90° C.
 8. Use of a cosmetic compositionwith an effective amount of extract from cistanche tubulosa (Schenk)Wight and other useful cosmetic adjuvant(s) in skin forfreckle-removing, whitening and/or improving the appearance of skin tobe youthful.
 9. (canceled)
 10. (canceled)
 11. The use according to claim8, wherein said cosmetic composition is applied to skin by the steps of:a. Applying a thin layer of the said composition on skin and leave it onindefinitely; b. Repeating the treatment 2 to 4 times daily.
 12. The useaccording to claim 11, wherein said cosmetic composition is applied toskin by further applying an occlusive patch or mask to skin aftertreatment.
 13. The use according to claim 8, wherein said cosmeticcomposition is applied to skin by the steps of: a) Providing saidcosmetic composition on a carrier of a nonwoven sheet or a hydrophilicgel patch comprising an extract from cistanche tubulosa (Schenk.) wightand at least one cosmetic ingredient; b) Applying the sheet or the patchto skin and leave it on for more than 10 minutes, and c) Removing thesheet or patch.